Pharmacological characterisation of capsaicin-induced relaxations in human and porcine isolated arteries

Capsaicin, a pungent constituent from red chilli peppers, activates sensory nerve fibres via transient receptor potential vanilloid receptors type 1 (TRPV1) to release neuropeptides like calcitonin gene-related peptide (CGRP) and substance P. Capsaicin-sensitive nerves are widely distributed in human and porcine vasculature. In this study, we examined the mechanism of capsaicin-induced relaxations, with special emphasis on the role of CGRP, using various pharmacological tools. Segments of human and porcine proximal and distal coronary arteries, as well as cranial arteries, were mounted in organ baths. Concentration response curves to capsaicin were constructed in the absence or presence of the CGRP receptor antagonist olcegepant (BIBN4096BS, 1 μM), the neurokinin NK1 receptor antagonist L-733060 (0.5 μM), the voltage-sensitive calcium channel blocker ruthenium red (100 μM), the TRPV1 receptor antagonist capsazepine (5 μM), the nitric oxide synthetase inhibitor Nω-nitro-l-arginine methyl ester HCl (l-NAME; 100 μM), the gap junction blocker 18α-glycyrrhetinic acid (10 μM), as well as the RhoA kinase inhibitor Y-27632 (1 μM). Further, we also used the K+ channel inhibitors 4-aminopyridine (1 mM), charybdotoxin (0.5 μM) + apamin (0.1 μM) and iberiotoxin (0.5 μM) + apamin (0.1 μM). The role of the endothelium was assessed by endothelial denudation in distal coronary artery segments. In distal coronary artery segments, we also measured levels of cyclic adenosine monophosphate (cAMP) after exposure to capsaicin, and in human segments, we also assessed the amount of CGRP released in the organ bath fluid after exposure to capsaicin. Capsaicin evoked concentration-dependent relaxant responses in precontracted arteries, but none of the above-mentioned inhibitors did affect these relaxations. There was no increase in the cAMP levels after exposure to capsaicin, unlike after (exogenously administered) α-CGRP. Interestingly, there were significant increases in CGRP levels after exposure to vehicle (ethanol) as well as capsaicin, although this did not induce relaxant responses. In conclusion, the capsaicin-induced relaxations of the human and porcine distal coronary arteries are not mediated by CGRP, NK1, NO, vanilloid receptors, voltage-sensitive calcium channels, K+ channels or cAMP-mediated mechanisms. Therefore, these relaxant responses to capsaicin are likely to be attributed to a non-specific, CGRP-independent mechanism.
Introduction
Capsaicin, a pungent constituent of red pepper, is known to activate sensory C-fibres via transient receptor potential vanilloid receptors type 1 (TRPV1; Caterina et al. 1997; Gunthorpe et al. 2002; Szallasi 2006), which are nonselective cation channels. Activation of these channels increases influx of mono- and divalent cations, which leads to an increase in the intracellular Ca2+ concentrations. Consequently, an array of neuropeptides like calcitonin gene-related peptide (CGRP), substance P and neurokinin A are released (Franco-Cereceda et al. 1988; Mitchell et al. 1995). These neuropeptides play a role in the regulation of normal vascular smooth muscle tone and are also implicated in several pathological conditions like ischemic preconditioning (Chai et al. 2006), preeclampsia (Dong et al. 2005) and migraine (Arulmani et al. 2004b). In the pathophysiology of migraine, vasodilatation of cranial blood vessels, especially extracerebral intracranial blood vessels, which are richly innervated by nerves containing a number of peptides such as CGRP, seems pivotal (Arulmani et al. 2006). Vasodilatation of these intracranial arteries leads to activation of nociceptors, which stimulate the pain centres in the brain (Goadsby et al. 2002). Involvement of CGRP in the pathophysiology of migraine is further strengthened by the observation that olcegepant (BIBN4096BS, Doods et al. 2000), a CGRP receptor antagonist, is effective in the acute treatment of migraine attacks (Olesen et al. 2004). Capsaicin has been widely used in various in vivo models of migraine to induce cranial vasodilatation, which is attributed to endogenous release of neuropeptides, especially CGRP (Akerman et al. 2003; Arulmani et al. 2004a; Gupta et al. 2006a). The involvement of CGRP is further substantiated in all these in vivo studies by the fact that the capsaicin-induced vascular responses are all amenable to blockade with CGRP receptor antagonists. Repeated capsaicin challenges are reported to deplete a number of neurotransmitters, including CGRP in various animal models (Tang et al. 1997; Tang et al. 1999; Zhou et al. 2002). In addition to the in vivo models, also in vitro vascular models involving meningeal (Gupta et al. 2006b) and coronary (Edvinsson et al. 2002; Gupta et al. 2006c) arteries have been used in migraine research, but vasorelaxation in these models was induced by exogenously administered CGRP. Several major vascular beds, including the meningeal and coronary (Franco-Cereceda 1988), are richly innervated with capsaicin-sensitive sensory fibres containing CGRP (Edvinsson et al. 1987). Meningeal blood vessels have been shown to be involved in the plasma protein extravasation in rat dura mater (Delepine and Aubineau 1997; Seabrook et al. 1996), a model of neurogenic inflammation, one of the putative mechanisms in migraine pathophysiology. In this model, capsaicin pre-treatment has been shown to block plasma protein extravasation, again by depleting neuropeptides (Delepine and Aubineau 1997). Therefore, it is of interest to investigate the release of endogenous CGRP in various blood vessels and capsaicin seems most appropriate for such studies.
Interestingly, very few studies have investigated capsaicin-induced relaxations in human isolated blood vessels (Franco-Cereceda 1991a; Franco-Cereceda et al. 1987). Based on CGRP-like immunoreactivity, it was claimed that capsaicin-induced relaxations are mediated by CGRP (Franco-Cereceda 1991b), but no CGRP receptor antagonists were used to unequivocally demonstrate the role of CGRP in these responses. Therefore, we were interested in studying capsaicin-mediated relaxations in human and porcine isolated arteries as a model to study endogenous release of CGRP in view of its relevance in the pathophysiology of migraine. Remarkably, some recent studies indicate that capsaicin-induced relaxations are mediated by non-CGRP-mediated mechanisms in guinea pig ileum, rabbit coronary arteries and equine tracheal smooth muscle preparations (Fujimoto et al. 2006; Yeon et al. 2001; Zhu et al. 1997), which is contrary to the observation that capsaicin-induced relaxations are mediated principally by CGRP. In these studies, relaxant responses to capsaicin were mainly attributed to different Ca2+-activated K+ channels. Therefore, in the present study, we tried to characterise capsaicin-induced relaxant response in human and porcine vessels, with special emphasis on the role of CGRP, and also investigated other mechanisms, using different pharmacological tools.
Materials and methods
Tissue preparation
The study protocol was approved by the ethical committee of Erasmus MC. Human coronary artery was obtained from ‘heart-beating’ organ donors (13 men, 19 women; 13–65 years) who died due to non-cardiac disorders. The hearts were provided by the Heart Valve Bank, Rotterdam, The Netherlands, after donor mediation by Bio Implant Services Foundation/Eurotransplant Foundation (Leiden, The Netherlands). The meningeal arteries were obtained from patients (four men, six women; 41–75 years) undergoing craniotomy at the neurosurgical unit (n = 6) or from the Department of Pathology (postmortem, n = 4) within 24 h of death, at Erasmus MC, Rotterdam. During the surgical procedure or autopsy, the dura mater together with a small piece of the meningeal artery was cut and placed in a plastic container filled with ice-cold (0–4°C), physiological salt solution. The artery segment was immediately transported to the laboratory and placed in cold oxygenated Krebs bicarbonate solution of the following composition (mM): NaCl 119, KCl 4.7, CaCl2 1.25, MgSO4 1.2, KH2PO4 1.2, NaHCO3 25 and glucose 11.1; pH 7.4. Excess tissue was removed. The meningeal artery (internal diameter of 350–800 μm) was used the same day or stored overnight in cold oxygenated modified Krebs solution and used the following day. Porcine hearts and heads (pigs of either sex; 6–12 months of age) were collected from a local slaughterhouse. Porcine basilar (internal diameter of 150–200 μm) as well as meningeal (internal diameter of 100–250 μm) arteries were dissected out from the skull and were placed in a cold oxygenated Krebs solution as described earlier for human meningeal arteries. In the laboratory, both human and porcine proximal (internal diameter, 2–3.5 mm) and distal (internal diameter, 200–600 μm) coronary arteries were dissected out of the right ventricle; distal coronary arteries were dissected with the aid of a microscope and stored in bicarbonate solution of following composition (mM): NaCl 118, KCl 4.7, CaCl2 2.5, MgSO4 1.2, KH2PO4 1.2, NaHCO3 25 and glucose 11.1; pH 7.4.
Functional studies
Both human and porcine proximal coronary artery segments (3–4 mm length) were suspended with the help of stainless steel hooks in 15-ml organ baths with a pretension of 15 mN (optimal tension shown in earlier experiments). Distal coronary and cranial artery segments were cut into ring segments of 1–2 mm length and were mounted in Mulvany myographs between two parallel titanium wires with a tension normalised to 90% of l100 (distance when transmural pressure equals 100 mmHg), thus achieving optimal conditions for active force development (Nyborg et al. 1987). Vessels were allowed to equilibrate for 30 min in Krebs solution at 37°C, a similar equilibration period was repeated after each physical or pharmacological challenge. In case of human arteries, the cyclo-oxygenase inhibitor indomethacin (0.1 μM) was added to prevent prostaglandin synthesis during the whole experimental protocol. Two successive challenges to KCl (30 mM) were performed to test the reproducibility of responses. Endothelial integrity was assessed by observing relaxation to substance P (10 nM) after precontraction with U46619 (9,11-dideoxy-11α, 9α-epoxymethano-prostaglandin F2α; 10 nM–1 μM), followed by washout of the agonists. KCl (18–30 mM, except where higher concentrations are indicated) or U46619 (10 nM–1 μM), in case of the K+ channel blockers, was used to obtain a stable contraction plateau of around 60–75% of the maximal contraction reached with KCl (100 mM). Subsequently, capsaicin was added in a cumulative manner in log steps. The different antagonists were used to antagonise capsaicin responses, and the concentration of each of these antagonists was decided on the basis of highest concentration in the literature unless mentioned otherwise. Concentration response curves were constructed in the absence or presence of the CGRP receptor antagonists olcegepant (1 μM; Gupta et al. 2006b,c) and CGRP8–37 (10 μM; Gupta et al. 2006b,c), the neurokinin NK1 receptor antagonist L-733060 (0.5 μM; Seabrook et al. 1996), the voltage-sensitive calcium channel blocker ruthenium red (100 μM, 45–90 mM KCl was required to get a similar precontraction as in the corresponding control segments; Franco-Cereceda and Rudehill 1989), the nitric oxide synthetase inhibitor Nω-nitro-l-arginine methyl ester HCl (l-NAME; 100 μM; Batenburg et al. 2004a,b), as well as the vanilloid receptor antagonist capsazepine (5 μM; Fujimoto et al. 2006; Gazzieri et al. 2006). At still higher concentrations of capsazepine (>5 μM) and L-733006 (>0.5 μM), even higher concentrations of preconstricting agents could not produce a workable precontraction.
We also investigated the role of different K+ channels by using various inhibitors, 4-aminopyridine (1 mM; Fujimoto et al. 2006; Yeon et al. 2001), a voltage-dependent K+ channel (Kv) blocker and charybdotoxin (0.5 μM), a blocker of Ca2+-dependent K+ channels for large conductance (BKCa) and intermediate conductance (IKCa), in combination with apamin (0.1 μM), a blocker of small-conductance Ca2+-dependent K+ channels (SKCa; Batenburg et al. 2004b; Fujimoto et al. 2006; Zhu et al. 1997). The effect of the RhoA kinase inhibitor, Y-27632 (1 μM; Fujimoto et al. 2006) was investigated both in the absence or presence of 4-aminopyridine (1 mM). We also investigated the effect of the gap junction inhibitor 18-α-glycyrrhetinic acid (10 μM) on capsaicin-induced relaxations. Where indicated, the endothelium was removed with a human hair, and removal was confirmed by observing a relaxation of less than 10% of precontraction of U46619 after addition of substance P.
Further, in porcine distal coronary artery, we also studied the effect of repeated administration of capsaicin (50 μM, four times) to verify the reproducibility of the responses in view of possible depletion of endogenous peptide pools or other agents.
Measurements of cAMP
Human proximal and distal, as well as porcine distal, coronary artery segments were incubated in a medium containing isobutylmethylxanthine (0.5 mM) for 30 min in the absence or presence of olcegepant (1 μM). The arterial segments were exposed to KCl (30 mM), challenged with ethanol (vehicle of capsaicin, final concentration in the baths 0.56%), capsaicin (10 μM) or h-αCGRP (100 nM, serving as a positive control) for 5 min and then snap frozen. The samples were stored at −80°C until cyclic adenosine monophosphate (cAMP) assay. To determine cAMP, tissues were homogenised in 0.5 ml 0.1 M HCl using a stainless steel ultraturrax (Polytron, Staufen, Germany). Homogenates were centrifuged at 3,300×g, and cAMP was measured in 300 μl supernatant using the ELISA kit according to the instructions of the manufacturer (R&D Systems Europe, Abingdom, UK).
Measurements of CGRP in organ bath fluid
Human distal coronary artery segments were subjected to a similar protocol as during the functional studies, while bath fluids were collected after construction of the concentration response curve. The bath fluids were collected from the segments treated with vehicle and capsaicin (100 μM), and Krebs solution was used as a control. Bath fluids were stored in tubes containing aprotinin (0.6 TIU/ml) and stored at −80°C. A competitive radioimmunoassay (Peninsula Lab INC., San Carlos, CA, USA) was used according to the instructions of the manufacturer to measure the CGRP concentrations in the bath fluid.
Data presentation and statistical analysis
The relaxant responses elicited by the agonists are expressed as percentage relaxation of the tone induced by 30 mM KCl or U46619 (in case of the K+ channel blockers). All data are presented as means±SEM, and n represents the number of blood vessel segments, all obtained from different subjects. The effect of all potential inhibitors of the relaxations to capsaicin was investigated in a paired parallel setup; that is, relaxations in segments with inhibitors were always compared to relaxations obtained in control segments from the same subject. The concentration response curves for all agonists were analysed using nonlinear regression analysis, and the potency of agonists was expressed as pEC50 using Graph Pad Prism 3.01 (Graph Pad Software, San Diego, CA, USA), setting the Emax of capsaicin in the presence of potential inhibitors to that in the respective control segment in case it was lower than the control Emax. The blocking potency of the antagonists was estimated by calculating median effective concentration (EC50) ratios, and apparent pKB values were calculated for the antagonists at each given concentration, with the slope set to unity. Statistical differences between concentration response curves to capsaicin in the absence and presence of potential inhibitors were determined using Student’s paired t-test with α set to 1.00–0.95(1/n) (Motulsky 2003) to correct for multiple comparisons. For the measurements of CGRP in the organ bath fluids, we could not exclude a non-Gaussian distribution due to the large degree of variability in the data. Therefore, the levels of CGRP in bath fluids were analysed by the non-parametric Kruskal–Wallis test, followed by Dunn’s post hoc multiple comparison test. Significance was assumed at P ≤ 0.05.
Compounds
Human α-CGRP and α-CGRP8–37 were obtained from Polypeptide, (Wolfenbüttel, Germany), olcegepant (BIBN4096BS, 1-piperidinecarboxamide, N-[2-[[5-amino-1-[[4-(4-pyridinyl)-1-piperazinyl]carbonyl] pentyl] amino]-1-[(3,5-dibromo-4-hydroxyphenyl) methyl]-2-oxoethyl]-4-(1,4-dihydro-2-oxo-3(2H)-quinazolinyl)-, [R-(R*,S*)]-) was a gift from Boehringer Ingelheim Pharma (Biberach/Riss, Germany); 4-aminopyridine was purchased from ICN Biomedicals (Aurora, OH, USA); L-733060 was purchased from Tocris (Bristol, UK); apamin, capsaicin, capsazepine, 3-isobutyl-1-methyl-xanthine, l-NAME, charybdotoxin, ruthenium red, substance P, U46619 and Y-27632 were purchased from Sigma-Aldrich (Zwijndrecht, The Netherlands), and KCl was obtained from Merck (Darmstadt, Germany). Capsaicin was dissolved in 70% ethanol, and the dilution series was also prepared in ethanol 70%. Capsazepine was dissolved in methanol; olcegepant was dissolved in a small amount of 1 N HCl and then diluted with distilled water. The other compounds were dissolved in distilled water, and all compounds were stored in aliquots at −80°C.
Results
Functional studies
Human arteries
Substance P relaxed artery segments precontracted with U46619 (10 nM–1 μM); responses were equi-efficacious in distal coronary (80 ± 5% of contraction to U46619, n = 28) and meningeal (75 ± 8%, n = 8) artery and significantly less in proximal coronary artery (27 ± 15%, n = 4). Both in the meningeal and distal coronary arteries, capsaicin induced concentration-dependent relaxations. In human proximal coronary artery, relaxant responses were only observed at the highest concentration of 100 μM, and the maximum relaxant response (34 ± 14% of contraction to 30 mM KCl) was significantly less than that observed in the distal arteries (94 ± 1% of contraction to 18–30 mM KCl). In human meningeal arteries, there was no difference in capsaicin-induced responses between arteries obtained perioperatively or postmortem; therefore, these data were pooled for further analysis. Capsaicin was equipotent and equi-efficacious in human distal coronary and human meningeal artery. In human distal coronary artery segments, the lower concentrations of capsaicin (0.1 nM–1 μM) in some cases induced contractions, but in all cases, we uniformly only measured the relaxant responses. The relaxations to capsaicin in proximal and distal coronary as well as meningeal arterial segments were insensitive to blockade by the CGRP antagonist olcegepant (1 μM; Fig. 1, Table 1).
Fig. 1Effect of capsaicin or its vehicle in the absence or presence of various pharmacological agents or interventions in precontracted human and porcine distal coronary arteriesTable 1Effect of various antagonists/interventions on capsaicin-induced relaxations in human isolated artery segmentsAntagonist or other intervention (n)Emax (%)Δ EmaxpEC50Δ pEC50Human distal coronary artery (Control) (32)94 ± 15.27 ± 0.12Olcegepant (1 μM) (10)89 ± 48 ± 44.84 ± 0.090.33 ± 0.16CGRP8–37 (10 μM) (5)96 ± 33 ± 44.79 ± 0.060.16 ± 0.08Capsazepine (5 μM) (13)91 ± 31 ± 25.10 ± 0.130.07 ± 0.16Ruthenium red (0.1 mM) (9)92 ± 35 ± 35.01 ± 0.130.21 ± 0.22L-733060 (5 μM) (7)94 ± 22 ± 26.03 ± 0.78−0.54 ± 0.34Denuded endothelium (5)90 ± 69 ± 55.34 ± 0.48−0.21 ± 0.58l-NAME (0.1 mM) (7)94 ± 23 ± 25.23 ± 0.48−0.24 ± 0.4518-α-Glycyrrhetinic acid (10 μM) (3)96 ± 20 ± 25.08 ± 0.28−0.15 ± 0.38Olcegepant (1 μM) + L-733060 (5 μM) (3)90 ± 61 ± 36.04 ± 0.230.06 ± 0.21 (Control) (10)97 ± 15.91 ± 0.324-Aminopyridine (1 mM) (6)95 ± 22 ± 35.64 ± 0.380.94 ± 0.46Charybdotoxin (0.5 μM) + apamin (0.1 μM) (8)97 ± 10 ± 15.80 ± 0.370.23 ± 0.37Iberiotoxin (0.5 μM) + apamin (0.1 μM) (5)96 ± 20 ± 25.94 ± 0.650.15 ± 0.38Y-276323 (1 μM) (3)99 ± 1−1 ± 25.39 ± 0.21−0.58 ± 0.14Y-276323 (1 μM) + 4-Aminopyridine (1 mM) (3)100 ± 0−3 ± 45.12 ± 0.180.33 ± 0.28Human proximal coronary artery (Control) (4)34 ± 144.30 ± 0.14Olcegepant (1 μM) (4)36 ± 164.40 ± 0.17Human meningeal artery (Control) (10)91 ± 5 5.04 ± 0.09Olcegepant (1 μM) (10)96 ± 1−5 ± 45.03 ± 0.07−0.07 ± 0.08Capsazepine (5 μM) (4)81 ± 98 ± 74.90 ± 0.310.11 ± 0.31Ruthenium red (0.1 mM) (3)74 ± 155 ± 145.13 ± 0.420.02 ± 0.62L-733060 (5 μM) (4)79 ± 128 ± 154.80 ± 0.31−0.09 ± 0.31Emax is the maximum relaxant response, expressed as percentage of the respective precontraction; pEC50 is the −logEC50, where EC50 is the concentration of agonist required to produce half the maximal response. The arteries were precontracted with KCl (18–30 mM) except where KCl (45–90 mM; bold) or U46619 (ital).
As the relaxant responses to capsaicin were small in human proximal coronary arteries and the availability of human meningeal arteries was very limited, further experiments were only carried out in human distal coronary artery. In this preparation, the CGRP receptor antagonist CGRP8–37 (10 μM), the TRPV1 receptor antagonist capsazepine (5 μM) and the NK1 receptor antagonist L-733006 (0.5 μM) also did not attenuate capsaicin-induced relaxations (Table 1). Similarly, there was no significant difference in relaxant responses in endothelium-intact or endothelium-denuded segments (Table 1). Also, in the absence or presence of the NO synthase inhibitor l-NAME (100 μM) or in the presence of the gap junction blocker 18-α-glycyrrhetinic acid, capsaicin caused equipotent relaxations compared to the respective control segments (Table 1). Various K+ channel blockers, namely, 4-aminopyridine (1 mM), charybdotoxin (0.5 μM) + apamin (0.1 μM) and iberiotoxin (0.1 μM) + apamin (0.1 μM) were also unable to block the relaxant responses to capsaicin. The RhoA kinase inhibitor Y-27632 (1 μM) alone or in combination with 4-aminopyridine was also unable to block the relaxant responses to capsaicin. Ruthenium red (100 μM), a nonselective blocker of Ca2+ transport through membrane channels, also did not significantly block these responses (Table 1). It is noteworthy that at higher concentrations, capsazepine and L-733060 significantly attenuated the responses to their respective preconstriction agents, and also in case of preincubation with Y-27632 and ruthenium red, 2- to 20-fold higher concentrations of precontracting agents were required to induce 60–75% of the maximal contraction reached with KCl (100 mM). At still higher concentrations of capsazepine (>5 μM) and L-733060 (>0.5 μM), even higher concentrations of preconstricting agents could not produce a workable precontraction. Additionally, the combination of olcegepant (1 μM) + L-733060 (0.5 μM) in human distal coronary artery or 4-aminopyridine (1 mM) + Y-27632 (1 μM) both in human and porcine distal coronary artery were also unable to block the responses to capsaicin. The vehicle of capsaicin (0.56% ethanol) did not induce any significant relaxations. It should be noted that the potency of capsaicin was higher on a precontraction with U46619 (10 nM–1 μM; pEC50 of capsaicin, 5.91 ± 0.32) than on a precontraction with KCl (18–30 mM; pEC50 of capsaicin, 5.27 ± 0.12; P = 0.029). However, irrespective of the precontracting agent used, the antagonists behaved in a similar fashion towards capsaicin. For example, the effects of 4-aminopyridine on capsaicin-induced relaxations were similar after precontraction with KCl (pEC50, 5.01 ± 0.05 and 5.12 ± 0.41; n = 7, in the absence and presence of 4-aminopyridine, respectively) and with U46619 (pEC50, 6.58 ± 0.47 and 5.64 ± 0.38; n = 6, in the absence and presence of 4-aminopyridine, respectively). In human meningeal artery, capsazepine, L-733006 and also ruthenium red did not block the responses to capsaicin (Table 1).
Porcine arteries
In porcine arteries, substance P induced relaxations in precontracted arteries to a varying degree in different vessels. The relaxations in distal coronary (64 ± 6% of contraction to U46619, 10 nM–1 μM, n = 26) basilar (44 ± 3%, n = 3) and meningeal (57 ± 15%, n = 3) artery were similar, whereas that in proximal coronary (8 ± 8%, n = 4) artery was significantly smaller. In porcine proximal and distal coronary arteries, capsaicin induced concentration-dependent relaxations. Unlike in human coronary arteries, the maximum response was not significantly different between porcine proximal and distal coronary arteries (Table 2). In both the arteries, olcegepant (1 μM) did not block the responses to capsaicin. Further experiments were carried out in distal coronary arteries and, similar as in the human distal coronary artery, capsazepine (1 μM), ruthenium red (0.1 mM), L-73360 (0.5 μM), 4-aminopyridine (1 mM), l-NAME (0.1 mM), charybdotoxin (0.5 μM) + apamin (0.1 μM), iberiotoxin (0.5 μM) + apamin (0.1 μM), Y-27632 (1 μM), 18-α-glycyrrhetinic acid (10 μM) and endothelium denudation did not affect capsaicin-induced relaxations (Fig. 1, Table 2). Additionally, Y-27632 (1 μM), alone or combined with 4-aminopyridine (1 mM), was also unable to block the responses to capsaicin. Similar as in the other arteries, the vehicle did not induce a relaxation (Fig. 1). Interestingly, in contrast to what we observed in the human distal coronary artery, precontracting the arteries with either KCl or U46619 did not change the relaxant responses to capsaicin. Four consecutive challenges to capsaicin (50 μM) did not significantly affect the magnitude of the responses (Fig. 2).
Table 2Effect of various antagonists/interventions on capsaicin-induced relaxations in porcine isolated artery segmentsAntagonist or other intervention (n)Emax (%)Δ EmaxpEC50Δ pEC50Porcine distal coronary artery (Control) (56)96 ± 15.27 ± 0.09Olcegepant (1 μM) (10)92 ± 23 ± 25.26 ± 0.11−0.02 ± 0.11CGRP8–37 (10 μM) (5)97 ± 3−2 ± 54.78 ± 0.060.00 ± 0.03Capsazepine (5 μM) (7) 99 ± 1−3 ± 25.15 ± 0.260.26 ± 0.16Ruthenium red (0.1 mm) (12)88 ± 37 ± 44.89 ± 0.090.47 ± 0.23L-733060 (5 μM) (7)88 ± 34 ± 44.89 ± 0.09−0.04 ± 0.15Denuded endothelium (9)91 ± 42 ± 25.62 ± 0.490.21 ± 0.16l-NAME (0.1 mM) (6)90 ± 88 ± 84.84 ± 0.150.28 ± 0.31 (Control) (15)99 ± 05.08 ± 0.1518-α-Glycyrrhetinic acid (10 μM) (3)99 ± 10 ± 15.73 ± 0.75−0.59 ± 0.534-Aminopyridine (1 mM) (7)96 ± 23 ± 24.84 ± 0.15−0.46 ± 0.35Charybdotoxin (0.5 μM) + apamin (0.1 μM) (11)99 ± 10 ± 15.27 ± 0.15−0.04 ± 0.20Y-276323 (1 μM) (9)96 ± 23 ± 25.17 ± 0.190.04 ± 0.21Y-276323 (1 μM) + 4-Aminopyridine (1 mM) (8)96 ± 34 ± 35.22 ± 0.200.00 ± 0.11Porcine proximal coronary artery (Control) (4)100 ± 05.33 ± 0.42Olcegepant (1 μM) (4)90 ± 65.79 ± 0.16Porcine basilar artery (Control) (3)97 ± 14.70 ± 0.05Olcegepant (1 μM) (3)100 ± 04.97 ± 0.24Capsazepine (5 μM) (3) 100 ± 04.80 ± 0.01Porcine meningeal artery (Control) (3)99 ± 14.82 ± 0.02Olcegepant 1 μM (3)99 ± 14.88 ± 0.04Emax is the maximum relaxant response, expressed as percentage of the respective precontraction; pEC50 is the −logEC50, where EC50 is the concentration of agonist required to produce half the maximal response. The arteries were precontracted with KCl (18–30 mM) except where KCl (45–90 mM; bold) or U46619 (ital).Fig. 2Effect of four consecutive challenges to capsaicin (50 μM) in porcine distal coronary arteries precontracted with KCl (30 mM)
There were no significant differences in efficacy or potency of capsaicin in porcine basilar (Emax, 97 ± 1%; pEC50, 4.70 ± 0.05; n = 3) and meningeal (Emax, 99 ± 1%; pEC50, 4.82 ± 0.02; n = 3) arteries as compared to porcine distal coronary arteries. In both these arteries, responses to capsaicin were not affected by olcegepant (1 μM).
Measurements of cAMP
Capsaicin (10 μM) did not affect cAMP levels in comparison to its vehicle or the control, which was only exposed to 30 mM KCl. In contrast, αCGRP (100 nM), which was used as a positive control, significantly increased cAMP levels in both human and porcine distal coronary arteries, which was blocked after incubation with olcegepant (1 μM; Fig. 3).
Fig. 3Changes in cAMP levels in human (n = 4–7) and porcine (n = 4–9) distal coronary artery segments after exposure to various pharmacological agents. *Significantly different (P < 0.05) from KCl (30 mM)-treated segments
Measurements of CGRP levels in organ bath fluid
Capsaicin and its vehicle induced a significant CGRP release from human distal coronary artery segments into the organ bath fluid (Fig. 4) as compared to levels observed in Krebs solution. There was no significant difference between CGRP levels obtained after incubation with capsaicin or its vehicle.
Fig. 4CGRP levels measured in bath fluids (Krebs buffer, control) after capsaicin or vehicle challenge in precontracted human distal coronary artery segments. *Significantly different (P < 0.05) from Krebs buffer
Discussion
In the present study, we investigated the role of CGRP in capsaicin-induced relaxations in human and porcine isolated arteries. In all arteries investigated, there does not seem to be any relevant role of CGRP in the relaxant responses to capsaicin. Further, the effects of capsaicin appear to be mediated by non-specific mechanisms.
In all arteries investigated, capsaicin induced concentration-dependent relaxations, although these had a limited potency and efficacy in human proximal coronary artery. The responses to capsaicin were resistant to blockade with olcegepant in all tissues studied, which suggests no involvement of CGRP receptors. The potency of capsaicin is in line with what has earlier been observed in human coronary arteries (Franco-Cereceda 1991a) and guinea pig ileum (Fujimoto et al. 2006). As expected in view of its resistance to olcegepant, CGRP8–37 (10 μM) also did not block capsaicin-induced relaxations in human distal coronary artery segments. This is contrary to the observations by Franco-Cereceda and Rudehill (1989), where the authors claim the relaxations to capsaicin are mediated by CGRP on the basis of the observation that the sustained relaxations in the human coronary induced by capsaicin are similar to those induced by exogenous CGRP, unlike the transient relaxations to substance P, which are followed by rapid tachyphylaxis. In another publication, these authors claim involvement of CGRP in responses to capsaicin in the human isolated coronary artery on the basis of increased CGRP-like immunoreactivity after exposure of the artery segments to capsaicin (Franco-Cereceda 1991b). It should be noted that in the above study, the authors show an increase in CGRP-like immunoreactivity in large arteries, where no functional studies were performed. Further, the classical prerequisite for demonstrating the involvement of a particular pharmacological agent, by using the corresponding antagonist to block functional responses, was lacking in this study. Admittedly, the same group did demonstrate that CGRP8–37 inhibited the responses to capsaicin in porcine coronary arteries (Franco-Cereceda 1991b), although we could not block these responses even at 10-μM concentration.
As CGRP-induced responses are mediated by increases in cAMP (Gupta et al. 2006c), we measured the levels of this second messenger after exposure to capsaicin and CGRP. We observed no increase in cAMP after addition of capsaicin, in contrast to the increased levels of cAMP after exposure of the vessel segments to (exogenous) α-CGRP. These increased levels were, as expected, blocked by the CGRP receptor antagonist olcegepant. Interestingly, after exposure to capsaicin, we observed CGRP release in the organ bath fluid, where the human distal coronary arteries were mounted in accordance with earlier observations (Franco-Cereceda 1991a). However, a similar increase was observed after administration of vehicle, while the control was not studied in the study of Franco-Cereceda et al. (1991a). Although capsaicin is known to activate TRPV1, there are reports that ethanol, also via activation of TRPV1, induces the release of CGRP as well (Trevisani et al. 2002, 2004). Moreover, as obvious from Fig. 1, the vehicle did not induce any relaxations in the precontracted arteries, and hence, the released CGRP cannot account for relaxations induced by capsaicin. Interestingly, the concentration of CGRP detected in the bath fluid of about 3 pM should have been about 10,000 times higher in the vessel segments (∼0.5 mg tissue in 5-ml organ bath fluid). Thus, the concentration of CGRP in the vessel segments should have been in the nanomolar range, which is equal to or even higher than the pEC50 in human distal coronary artery under similar experimental conditions (Gupta et al. 2006c) and should, thus, have induced a detectable relaxation. Therefore, it is most likely that the radioimmunoassay displayed cross-reactivity to another ligand, not CGRP. Admittedly, our observation that ruthenium red, even at a very high concentration (100 μM), did not block the responses to capsaicin, is in contrast with the observations described by Franco-Cereceda (1991a), where ruthenium red completely blocked the responses to capsaicin. Additional evidence for the fact that CGRP and TRPV1 are not involved in relaxant responses to capsaicin is provided by the fact that capsazepine, a competitive antagonist of TRPV1 (Caterina et al. 1997), did not block responses to capsaicin in our study.
In view of reports of involvement of various K+ channels in the relaxant responses to capsaicin in various smooth muscle preparations (Yeon et al. 2001; Zhu et al. 1997), we also investigated various K+ channel inhibitors in similar or even higher concentrations, but none of these blocked the responses to capsaicin. 4-Aminopyridine, a blocker of delayed rectifier K+ channels, which is reported to block the responses to capsaicin in rabbit coronary artery (Yeon et al. 2001) and guinea pig ileum (Fujimoto et al. 2006), was ineffective in human as well as porcine distal coronary artery. Similarly, charybdotoxin, a BKCa and IKCa blocker, which blocked capsaicin responses in equine tracheal smooth muscle (Zhu et al. 1997), did not block these responses at similar concentrations and even in combination with apamin, a SKCa blocker. We also used the RhoA kinase inhibitor Y-27632, but unlike in guinea pig ileum, (Fujimoto et al. 2006) it also did not antagonise responses to capsaicin in the isolated preparations used in the present study. The capsaicin-mediated responses appear to be endothelium-independent, as denudation of the endothelium did not significantly change the capsaicin-induced responses. Further, the vehicle of capsaicin was without effect and, hence, cannot account for the relaxant responses to capsaicin. Moreover, repeated administration of capsaicin in porcine coronary artery did not significantly decrease the responses, suggesting that stored neuropeptides are not responsible for the relaxations, as these would most likely be depleted after repeated challenges to capsaicin (Sams-Nielsen et al. 2001). Unlike in the present study, relaxations in precontracted guinea pig isolated pulmonary artery could not be repeated with capsaicin indicating depletion of neurotransmitters (Maggi et al. 1990). Although we did not include a positive control for the various K+ blockers in the current study, in the same set up at our laboratory, relaxant responses to l-S-nitrosocysteine were blocked by the combination of charybdotoxin (100 nM) and apamin (100 nM) in porcine distal arteries (Batenburg et al. 2004b). Further, the concentrations of the various inhibitors that we used in our study were equal to (Yeon et al. 2001; Zhu et al. 1997) or even higher (Yeon et al. 2001) than those employed by others.
Taken together, our observations in human and porcine distal coronary artery suggest that capsaicin-induced responses are not mediated by CGRP, substance P or TRPV1 receptors and also do not involve various Ca2+-activated K+ channels. The relaxations to capsaicin are mediated by a cAMP independent pathway. The major component of capsaicin-induced relaxations, therefore, appears to be mediated by non-specific actions of capsaicin, rather than from the release of neuropeptides like CGRP.

capsaicin
cgrp
human coronary artery
human meningeal artery
porcine coronary artery


Using Soil and Water Conservation Contests for Extension: Experiences from the Bolivian Mountain Valleys

Soil and water conservation (SWC) contests among farmer groups were organized in five rural villages in the Bolivian mountain valleys. The contests were aimed at quickly achieving widespread sustainable results. This article analyzes the effectiveness of these contests as an extension tool. Mixed results were obtained. In three villages, participation rates in the SWC activities introduced in the contests were still high even 2 years after project withdrawal. These were all villages where a solid foundation for sustainable development had been laid before the contests were held. Two years later, most families were still involved in maintenance of the SWC practices introduced in the contests, and many farmers had started to experiment with different soil management practices. However, replications of these SWC practices were not widespread, Conservation Leaders did not continue with their training activities, and the quality of maintenance of the practices was often not satisfactory. In order to become a more effective extension tool and achieve widespread impact, SWC contests must receive continued support by a catalyst agency. Moreover, other SWC contests should also be organized in which practices are not predefined. Given that SWC contests are a low-budget extension tool, local municipalities could become more actively involved.
Introduction
Agricultural development programs and extension services, both in developed and developing countries, struggle with frustratingly low adoption rates of soil and water conservation (SWC) practices. In response, Savenije and Huijsman (1991) called for a “making haste slowly” approach that focuses more emphasis on developing and implementing solutions together with rural people. Numerous experiences, principally from Non-Governmental Organizations (NGOs), have proven that these more participatory (slow) approaches often work and that sustainable livelihoods can be established. Successes to date, however, consist of scattered small-scale projects; wide-scale impact (haste) has yet to be achieved. The major dilemma remains combining widespread and short term impact (Farrington 1998). In SWC extension in particular, participatory approaches are too limited in scope, while existing governmental extension services are often not participatory enough and not sustainable. Hence, strategies and accompanying methodologies and tools are needed that achieve sustainable results in a short time and on a widespread scale.
Despite urgent calls to modify attitudes and develop strategies for the extension of SWC (Sombatpanit and others 1996), the agricultural extension service in Bolivia has completely disappeared, and a governmental strategy for tackling soil erosion does not exist. NGOs have partially filled this gap, and several small-scale successes were obtained with the participatory introduction of SWC practices. However, these successes have not been reproduced at a large enough scale to have an impact at national (or even departmental) level. Government officials are often not even aware of these experiences, and NGOs themselves generally do not have the capacity for wide-scale dissemination of technology options (Farrington 1998). As a consequence, land degradation during the past decades has increased (Kessler and Stroosnijder 2006), and rural poverty remains a serious threat to sustainable development in Bolivia.
In response, a project executed by the Japan Green Resources Corporation (JGRC) developed and validated a strategy to motivate poor Bolivian mountain farmers to engage in adequate natural resources management, the logical strategy for SWC (Kessler 2007a). Unlike other small-scale participatory experiences, the logical strategy not only aims at slowness but also at making haste (i.e., in finding participatory methodologies and tools that achieve sustainable widespread success in SWC). Scaling-up and integrating SWC in a future extension system at a national level is the ultimate objective. One of the logical strategy’s most innovative tools is conducting SWC contests among organized groups of farmers. The objective of this article is to analyze the effectiveness of these SWC contests as an extension tool. This is principally done by monitoring and evaluating farmers’ participation in maintaining and replicating SWC practices after execution of the contests. The results will be used to discuss possibilities of using SWC contests in an extension strategy at a national level.
Study Area
The study was conducted in the five research villages of the JGRC project. They are situated in the north-Chuquisaca region of the inter-Andean valleys of Bolivia (Fig. 1), which is a semi-arid region located at an altitude of 2500 to 3100 m above sea level. The majority of the families in this region still manage a mixed farming system, with mainly subsistence agriculture and a flock of goats and sheep. Potatoes are still the main crop and are the only crop that receives small amounts of manure. Other important crops are maize, wheat, and barley. Animal traction is mostly used for land preparation. Chuquisaca is one of the least developed areas in Bolivia, with the second lowest Human Development Index (HDI) in Bolivia, equaling only 0.49 (United Nations Development Programme 1997). Similarly, a study by the JGRC project revealed that the Human Poverty Index (HPI) for the research villages is among the highest in the world (on average 45%), with lack of access to drinking water and health facilities being the principal constraints. Hence, the region is very poor and faces enormous socioeconomic and physical constraints.
Fig. 1Location of the study area in the inter-Andean valleys of Bolivia
In three of the research villages, the “experimental villages” (Tomoroco, Kaynakas, and Sirichaca), the logical strategy was developed and tried out over a period of 4 years. The other two villages were “validation villages” (Talahuanca and Patallajta); in these villages the strategy was validated for 2 years. The villages were carefully selected, in order to obtain a representative sample of villages for the north-Chuquisaca region. Therefore, geographical, climatic, cultural, and socioeconomic characteristics are different among the villages. Rainfall varies from 350 mm/year (in Tomoroco) to 750 mm/year (in Kaynakas). Sirichaca has more flat lands for potato production, whereas the other villages have steeper and more erosion-prone slopes.
SWC is a barely developed activity in this region. Since the colonial era, when farmers started using a combination of Spanish and traditional agricultural practices to maintain adequate production levels, practices have not changed much. As a consequence, once cultivable land began to be used more intensively, traditional conservation practices were no longer able to control erosion and maintain soil fertility. Most of the SWC practices currently found in the research villages are still based on traditional knowledge, but given their ineffectiveness in conserving soil and water, farmers have become increasingly sceptical about prospects for sustainable agriculture in their villages. Therefore, migration rates are high in most of the villages. Attempts of development agencies to promote improved SWC practices have not been successful, mainly because of failing intervention strategies and the lack of adequate extension tools.
Research Methodology
The SWC contests were conducted in the “experimental villages” in 2001, and in the “validation villages” in 2003. In each ‘‘experimental village’’ only one SWC contest, which focused on all physical SWC practices, was conducted; in the “validation villages” two SWC contests were conducted, focusing on two SWC practices each. During the SWC contests, the number of labor days invested per family was monitored for each SWC practice. Moreover, the rate of participation in the contests (i.e., percentage of families actively involved in SWC activities) was evaluated in each village.
The most important data for this study were obtained in the ex-post evaluation in 2005, 2 years after the JGRC project’s withdrawal from the villages. The ex-post evaluation was conducted in all villages, except Sirichaca. In this village an evaluation was not considered necessary, due to the disappointing results and low participation rates during the contests. In the other villages, a random sample of 30 families was taken for the ex-post evaluation. Although the diversity of farmers within each village is high, and farmers who are progress-driven and economically better-off tend to invest more in SWC practices than others (Kessler 2006), this was not considered a variable. Hence, large families, as well as, for example, widows were included in the sample. Similarly, farmers from the higher economic stratum (with more land) as well as very poor small farmers were considered.
In the ex-post evaluation, the percentage of families actively involved in both maintenance and replications of SWC practices was assessed by means of a field survey. If at least one type of SWC practice was properly maintained, the family was considered to be actively involved in maintenance. If at least one SWC practice was replicated on other fields after the project’s withdrawal, the family was considered to be actively involved in replications. Similarly, the active usage of other (nonphysical) SWC practices was evaluated during the ex-post evaluation.
Finally, in order to obtain data concerning the popularity of the executed SWC practices, quality of maintenance of these practices was evaluated within the same sample of families. For each family an average score was given for the quality of maintenance of each type of SWC practice, ranging from very bad (or abandoned) to very good. Based on these scores, and in order to be able to compare the villages, average scores for each village were calculated.
The Need for More Effective Extension Tools
Although Bolivia has tried out different extension service models (Bojanic 2001), these were always technology-centered and top-down, with weak research-extension linkages. Most widely used was the Training & Visit extension system, in which knowledge trickled down from the research institutes to the extension worker and finally to the farmers. Farmer participation in this model was mainly functional. In 1991 a World Bank project strengthened the country’s research capacity, but extension remained extremely weak, resulting in poor adoption rates of improved practices (Bojanic 2001). In contrast to other developing countries, where low adoption rates forced extension agencies to apply more people-centered extension approaches, the governmental extension service in Bolivia simply disappeared. The research and extension component remained an independent foundation only for the study of potatoes. Currently, a demand-driven research and extension system (SIBTA, the Bolivian Agricultural Technology System) is in place that promotes innovative technologies in support of productive chains. Given the system’s focus on promoting regional cash-crops, (poor) subsistence farmers are largely excluded.
In Bolivia, as in many other Latin America countries, NGOs fulfilled the more intensive extension tasks. In contrast to the functional type of participation of public sector organizations, most NGOs aim at an empowering type of participation (Farrington 1998). Several NGO-initiated experiments with Farmer Participatory Research (FPR), were based on the farmer-first approach (Chambers and others 1989). Their pioneering work with a large set of participatory techniques and innovative extension approaches has led to the realization that community participation and integrated (multi-sector) approaches are essential elements for success. In Latin America, farmer-to-farmer extension has had a notable impact and has led to adequate natural resources management in Mexico (Ramos 1998), Honduras (Sherwood and Larrea 2001), and Nicaragua (Braun and Hocdé 2000), among other countries. Given that NGOs can spend considerable resources in a few villages, and often invest in time-demanding and costly face-to-face participation, they have achieved higher participation rates than governmental extension services. However, the principal constraint of these extension approaches is their limited scope: they are difficult to replicate on a wide scale in the absence of local support organizations (Farrington and others 2002). With the prime objective of agricultural extension being to reach all farmers, this is a crucial constraint.
The JGRC project’s logical strategy for SWC is not the first attempt to develop effective strategies that combine farmer participation at the grassroots level with extension and scaling-up. Killough (2005), for example, proposes “participatory extension through [the] accompaniment model.” This extension approach is a middle road between the traditional extensionist-centered approaches and the more recent farmer-led approaches. In this model, professional extension workers help farmer-promoters conduct on-farm experiments and provide farmer-to-farmer training. Local empowerment (through the development of farmer leaders) and strengthening local institutions are primary goals of this holistic approach. It is therefore important that such extension approaches promote integrated human development. Evidence from Honduras shows that farmers would have likely abandoned SWC practices long ago, if the extension approach had not addressed essential aspects of the interaction between human development and agriculture (Sherwood and Larrea 2001). Farmers conceptualized this interaction and holistic approach as “the human farm.”
Implementing such an integrated and farmer-based extension approach is urgently needed in Bolivia; only then can a wide-scale impact be achieved. Although municipalities have considerable budgets for rural development, they do not have the human capacity to provide this service, and they regard extension as a state responsibility. Hence, there is a need for (1) profound changes at institutional level to give priority to extension, and (2) effective extension tools that can be used in a farmer-based extension approach. SWC contests are an example of such a tool.
Using SWC Contests Within a Farmer-Based Extension Approach
Organizations in several countries have experimented with farmer contests or competitions. The State Farm Bureaus in the United States regularly hold such contests, in particular for young farmers. Management, growth and progress in farming operations are major factors in judging the contests. The outstanding farmers receive financial awards, which are usually made available by sponsors. Farmer competitions in New Zealand and Australia are more focused on developing technical skills and knowledge, with conservation and land management practices being one of the several competition modules. In developing countries all kinds of farmer contests are held, but rarely focused on SWC. Chuma and Murwira (1999) report of farmer competitions in Zimbabwe organized to stimulate the process of experimentation and revival of farmer knowledge regarding best farming practices.
The best results with respect to SWC have been achieved in two rural development projects in Bolivia and Peru, where SWC contests among farmers were used to speed up execution of SWC practices. Both projects make use of the “learning from the best” principle (i.e., learning from the best families and villages) (Van Immerzeel and De Zutter 2005). Knowledge management (i.e., combining knowledge and capacities from different farmers to find sustainable and fast solutions) is essential in this approach. The contests are used as a catalyst tool to disseminate this knowledge and motivate farmers to learn from the best, to experiment and innovate, and to win prizes by improving on what they have learned (Van Immerzeel and De Zutter 2005).
In Peru, the rural development project, MARENASS (Management of Natural Resources in the Southern Highlands), uses farmer contests to promote new technological practices for improving natural resources management, agricultural production and living conditions. An important characteristic of the project is the transfer of decision-making and responsibility for planning and financial resources to the villages. Each participating village receives financial support to hire direct technical assistance (Posthumus 2005). These external service providers can be farmers with much experience (“the best farmers”), consultants or technical staff members. When a contest is organized, the villages themselves select and contract these privatized services to provide training to a number of selected farmers. By means of farmer-to-farmer training, the trained farmers in turn teach the other villagers the new techniques they have learned. Contests are held both at village level, with farmers competing against each other, and on district level between villages (Posthumus 2005). Jury members are selected by the participants; the families, or villages that best apply the recommendations provided by technical staff earn a cash prize. The innovations of MARENASS have been successful, and the project is still in progress (International Fund for Agricultural Development 2005); in 360 villages about 60% of all households have been reached (De Zutter 2004). Nevertheless, since internal problems in some villages limit their participation in the project, the impact of activities on watershed level is quite low (Posthumus 2005).
In Bolivia, the SID (Strategies for International Development) project, Pachamama Urupa, approaches the dual problems of soil erosion and rural poverty with the understanding that neither can be resolved without simultaneously addressing both. Competitions among villages are organized to encourage participation in natural resources management, and to recognize the most successful farmers (Strategies for International Development 2005). SID hires farmers who are skilled in one or more of the land conservation and reclamation practices as part-time staff. These para-professional extension agents train selected farmers in about five villages, who in turn train all the other participants of the contest in their respective villages (Borda 2002). The competitions are flexible enough to permit farmers to find their own solutions to their problems (i.e., to experiment and innovate). All farmers participate in the judging, which stimulates the sharing of knowledge and the adoption and improvement of the practices. The winning villages and families receive farm tools, seeds, and animals as prizes. Participation rates in the competitions are about 80% to 90%. The project has recently won the World Bank’s “Development Marketplace Competition” for innovative ideas in international development.
The MARENASS and SID projects are quite similar; they aim at wide-scale adequate natural resources management through a farmer-based extension approach that builds on local knowledge and capacities. Massive participation in farmer contests and farmer-to-farmer training are crucial aspects of this approach. Moreover, progressive learning and improvement is stimulated through constant interchanges, the participatory judging process and the closing ceremonies. Two important differences between the projects are that:in Bolivia the practices being judged in a contest are specifically defined, while in Peru the contests have a more general character (e.g., soil conservation) and practices are not specified;in Bolivia money is not involved (and prizes are in the form of goods), while in Peru farmers are responsible for contracting the trainers and cash prizes can be won.Based on these experiences, SWC contests are undeniably a very promising tool for farmer-based extension strategies, and for achieving fast and widespread sustainable impact in natural resources management. In the following section we will focus on the SWC contests of the JGRC project.
SWC Contests in the JGRC Project
In the logical strategy of the JGRC project, SWC practices are executed within a framework of integrated rural development. They are always accompanied by – and often integrated in – other activities that aim at improving local livelihoods. An important feature of this strategy is the laying of a solid foundation for sustainable development before starting a SWC contest, with activities focusing, for example, on better village organization, responsible participation and effective collaboration (Kessler 2007b). The project’s strategy stresses the human dimension of sustainable development: genuine participation of stakeholders is essential (Kessler 2007a). The objective of the SWC contests is to train farmers in basic SWC practices. Moreover, the contests encourage farmers to experiment and to decide which practices best fit their specific conditions.
Reflection and dialog – two key features of Participatory Research & Extension (Percy 2005) – are constantly used. Participatory research with a selected group of farmers, as well as farmer-to-farmer training and knowledge transfer, are crucial before, during and after the SWC contests. Lessons learned through the farmer-to-farmer movement that began in the Guatemalan highlands (Bunch 1982) are, therefore, taken into account (e.g., to use small-scale experimentation) to start slowly and small, to achieve early recognizable success and to limit the introduction of technology. The essential multiplier effect is provided by the SWC contests and farmer-to-farmer training.
In the next section we explain the SWC contests in more detail. In all the activities the project’s extension worker plays a crucial role. At the end of the section we also present the differences between the approach of the aforementioned two projects and the JGRC project.
Activities Preceding the SWC Contests
Preceding the SWC contests (i.e., during the laying of a solid foundation for sustainable development in a village), SWC activities start with a group of about 10 Conservation Leaders (CLs). CLs are chosen by the assembly, taking into account personal characteristics such as responsibility, honesty and willingness to innovate. They receive intensive training from the project’s extension worker, aiming at the generation of a progress-driven attitude and at conducting experiments on their fields. Experimentation focuses both on physical SWC practices (stone lines, diversion ditches, bench terraces, etc.) and agronomical soil management practices (more efficient manure use, composting, green manure, etc.). CLs are also stimulated to establish some test-sites for comparing with and without cases, with the objective of obtaining more visible results and making on-site comparisons. This might convince visiting farmers that the positive effects are indeed a result of the practices, and not of different physical conditions between their farm and the CLs’ farms. Apart from experimentation and providing demonstrations on their own farms, CLs also have many less tangible tasks such as mobilizing the villagers to become involved in development activities. They are thus both promoters and technicians. Finally, training of CLs in techniques for knowledge transfer is given before and during the farm visits.
Once a solid foundation is laid in the village and CLs are sufficiently trained, the group decides when the first SWC contest will be held and which practices will be executed. In the dry season, practices like stone lines and gully control measures are considered and in the wet season practices that require digging such as diversion ditches and bench terraces. The village is informed in the assembly and by distributing information leaflets. During a period of at least a few weeks, families have the opportunity to decide whether to participate in the contest. CLs have an active role in motivating their neighbors, and in starting to organize groups based on vicinity. Eventually, each CL should lead a group of five to eight families. During these weeks of group formation, possible conflicts must be resolved, especially between neighbors. Given that effective group collaboration is crucial for a successful SWC contest, group formation should be given the required time.
Execution and Evaluation of the SWC Contests
A contest generally deals with two SWC practices. In a later stage a certain contest can be repeated and/or more integrated contests can be considered, combining different practices. The first contest is the most important one, because it serves as a selection tool for distinguishing the interested families (those with a progress-driven attitude) from the others. In this first contest a subsidy is given – as an incentive to all participants – for the purchase of a set of tools that are essential to conduct SWC practices. Each family pays 20% of the original cost. Families that start participating in a later stage have no access to the subsidized tools, and should themselves catch up with the already executed practices.
Each SWC contest takes about one or two months. Twice a year a contest can be held; one in the dry season and the other one in the wet season. The criteria for evaluating the contests are clearly indicated before starting: (1) executed quantity, (2) quality of the work, (3) knowledge, and (4) group collaboration. Major emphasis is given to training and learning during the contests. An essential technique for conducting most of the SWC practices is the adequate handling of the A-frame, which is used for establishing the contour lines in a field. This is generally taught during the first contest.
Once the contest starts, each group – under the guidance of their CL – decides how the work will be done. Reciprocal group work (or “ayni” in Quechua) is mostly used. In this system, farmers work on each others’ farms on a rotation basis. It is especially useful for labor-intensive work, and hastens the pace of execution. The host family provides food; money is never used. Reciprocity in the Andes region is based on mutual trust. Torrico and others (1994) argue that it still contains many religious aspects, and that, therefore, quality of the work is never discussed. Similar to the “alayon”, a traditional form of village cooperation in the Philippines (Moneva and others 2000), the ayni serves as a venue for group learning, problem solving and the promotion of equitability among farmers. SWC practices are mainly conducted on fields situated near the farmer’s homestead; later they can be replicated on other fields. The project’s extension worker regularly monitors group work, and assists the CLs in their training. In the beginning, CLs often encounter problems and feel uncertain; regular meetings help to solve this.
Once a contest has finished, each CL measures the quantity of practices executed by his or her group. The verification of this quantity, as well as the evaluation of quality and knowledge, is done by means of cross-visits with other CLs. Quality criteria are harmonized between CLs before starting the evaluation. Knowledge is evaluated by asking some practical questions. For these three criteria (quantity, quality and knowledge) scores of 1 (bad) to 3 (good) can be obtained. This is written down on evaluation sheets. Based on observations, the extension worker evaluates the fourth criterion: group collaboration and cohesion. All criteria are given the same weight-factor in the final calculation.
During the final closing ceremony the groups receive a reward for their efforts. Recognition of efforts is important because it engenders a sense of pride and it increases self-confidence (Cinnéide and Conghaile 1990). The most recommendable rewards are seeds for green manure and vegetables, which contribute to more sustainable agriculture. Additionally, the winning groups also receive, for instance, potato, maize, or barley seeds, all in small quantities. These prizes are useful products and their value is small enough to avoid participating in the SWC contests for the wrong reason. In this respect, money or food must never be used as prizes.
Activities Succeeding the SWC Contests
Maintenance of the newly constructed practices is the first priority, and complementary vegetative and soil management practices are essential in order to achieve impact on soil productivity. Vegetative conservation practices (grasses, bushes, or trees) also strengthen most SWC practices and make them more sustainable. They require, however, controlled grazing and strict rules at village level that are respected by all villagers. Hence, only when such regulations are collectively agreed upon will vegetative practices be viable and will SWC practices work.
Given the importance of vegetation, establishing tree nurseries in each village – preferably at family level – is part of the holistic approach of the logical strategy. By means of an Integrated Project (Kessler 2007a), a group of farmers interested in agroforestry is trained to become trainers in this topic. Through farmer-to-farmer training, technical knowledge regarding agroforestry practices can be spread to a large number of farmers. Similarly, other groups of farmers specialize in, for example, manure management (improved stables, manure storage and collection methods), composting or green manure practices.
The spreading and replication of SWC practices to all the other fields that need to be conserved is the responsibility of each family. Although, ideally, groups that participated in the SWC contest will continue to work in ayni, most families will have to do it on their own. The role of the CLs is to provide support whenever it is requested.
Differences with the MARENASS and SID Projects
Although similar in many aspects, especially in considering SWC as being part of an integrated approach to sustainable rural development, the JGRC project conducted its SWC contests in a slightly different way than the MARENASS and SID projects. This concerned four crucial aspects:The emphasis on training in specific SWC practices during each contest instead of leaving more space for experimentation. The reason: the SWC contests are foremost an extension tool, i.e., they aim at providing farmers with basic practical knowledge of some simple SWC alternatives. After the contests farmers experiment with innovations and will adapt and improve the practices.The emphasis on laying a solid foundation for sustainable development before starting the SWC contests. The reason: only farmers with a progress-driven attitude will continue to experiment and innovate after the project’s withdrawal.The use of contests between groups instead of families or villages. The reason: to stimulate group formation and collaboration within a village, i.e., to maintain or re-establish the traditional work in ayni, and to contribute to better internal relations and knowledge exchange.The subsidizing of tools to stimulate participation in the contests, but no (or insignificant) prizes for winning groups. The reason: not having the tools is often a major limitation for participating, but once farmers participate, they must become convinced by the result of their work and not by the prizes they can win.
Results and Discussion
Table 1 shows that similar numbers of family labor days were invested during the SWC contests in all villages. Groups generally worked two aynis (or two complete labor days) on each group member’s fields. Stone lines were the most popular practice executed during the contests, with labor accounting for 40% to 70% of total labor days invested. In Tomoroco and Kaynakas – with more steeply sloping land – considerable investments were made in bench terraces, while in Sirichaca and Patallajta gully control works (especially the smaller ones) and earth bunds (due to the absence of stones) were given more attention.
Table 1Average number of labour days invested per family for each SWC practice, during the SWC contestsSWC practicesExperimental villagesValidation villagesTomorocoKaynakasSirichacaTalahuancaPatallajtaStone lines 8.15.64.55.25.4Diversion ditches 0.80.81.22.82.8Gully control0.30.61.80.52.5Bench terraces 2.12.7–––Earth bunds –0.11.60.21.1Infiltration ditches 0.20.3–––Individual terraces–0.51.4––Total 11.510.710.58.911.8Source: M&E data in 2001 (experimental villages) and 2003 (validation villages)
Table 2 shows that the percentage of families participating in the construction of practices during the SWC contests was lowest in Sirichaca (48%) and Patallajta (66%); these are both villages in which a solid foundation for sustainable development was never laid. In Tomoroco the participation rate was highest with 86%. These data show that in the villages where activities concerning organization, collaboration and environmental awareness raising had already been successfully executed, the SWC contests were able to mobilize more people.
Table 2Percentage of families actively involved in SWC activitiesSWC activityExperimental villagesValidation villagesTomorocoKaynakasSirichacaTalahuancaPatallajtaConstruction of SWC practices during the SWC contests86754884 66Maintenance of SWC practices two years after project withdrawal9169–8453Replications of SWC practices two years after project withdrawal78252030Source: M&E data in 2001 and 2003 (during the contests) and ex-post evaluation data in 2005
However, initial motivation is easy; continued motivation is what really matters (Savenije and Huijsman 1991). The effectiveness of the SWC contests can only be properly measured by evaluating the continued use of SWC practices after the project’s withdrawal. Table 2 shows the results of the ex-post evaluation, in which the percentage of families that have maintained and replicated SWC practices was assessed.
Concerning maintenance, in Tomoroco this percentage is highest and has even increased; presently, 91% of all families perform adequate maintenance of one or more SWC practices. Quality of maintenance was moderate to good in Tomoroco (Table 3); stone lines and bench terraces were especially well maintained. In Kaynakas 6% fewer families are currently involved in SWC activities; some families have not maintained their practices or have even removed stone lines and gully control measures. In some cases stones from stone lines were used for fruit tree terraces; such farmers experimented with and adapted practices. Farmers often refine their practices under environmental pressure (Veihe 2000). They may, for example, consider stones more effective for terrace building than for stone bunds. This is supported by Table 3: in Kaynakas the quality of maintenance is better for bench terraces than for stone lines. Bench terraces are popular and productive for vegetables.
Table 3Quality of maintenance of SWC practices two years after project withdrawalSWC practicesExperimental villagesValidation villagesTomorocoKaynakasTalahuancaPatallajtaStone lines++/−+++/−Diversion ditches+/−+/−+-Gully control+/−-+/−+/−Bench terraces++n.a.n.a.Earth bundsn.a.n.a.––++ very good; + good; +/− moderate; - bad; – very bad (abandoned); n.a. not applicable (not executed)Source: Ex-post evaluation data in 2005
Abandonment of SWC practices is highest in Patallajta, where two consecutive severe drought years and lack of impact of SWC practices caused general disillusion among the villagers. Migration increased and fields were left unattended. Table 3 shows that quality of maintenance is bad to moderate in this village; earth bunds, which were heavily damaged during a high-intensity rain storm, were all abandoned. In neighboring Talahuanca, however, despite severe drought and similar damage to earth bunds, all participating families in the SWC contests are still actively involved in maintenance, except for earth bunds. The presence in Talahuanca of a solid foundation for sustainable development before starting the SWC contests explains the differences between both ‘validation’ villages. Maintenance of gully control measures was generally given little attention in all of the villages, although most people are of the opinion that these practices work very well.
Concerning replications, a field survey in 2003 revealed that in Tomoroco and Kaynakas wide-scale replications were conducted during the two years in which the JGRC project was in effect. All participating families in the SWC contests constructed new SWC practices. This was mainly attributed to the inclusion of SWC activities in other activities of the JGRC project (namely Integrated Projects, see Kessler 2007a). Stone lines and diversion ditches were mostly replicated and covered large areas of the agricultural land in both villages. The average investment in these villages in maintenance and replications of SWC practices was estimated to be 20 labor days per family during these two years (2001 to 2003).
However, more important from a sustainability viewpoint is what happened after the project’s withdrawal (starting in 2003). Table 2 shows that in Tomoroco 78% of all families constructed replications of SWC practices in the subsequent two years; in the other villages this was much lower (about 25%). Stone lines, especially, were replicated, given their relatively low labor requirements. The differences between the villages are explained by the fact that Tomoroco has more potential land for constructing stone lines, while in Talahuanca and Kaynakas the most important fields had already been protected. Moreover, on the steeper slopes of Kaynakas, stone lines sometimes disturb land preparation. In this village bench terraces were found more useful, but their replication requires higher investments. Replications of stone lines were found in only one village outside the project area, near Tomoroco..
Hence, the SWC contests have achieved mixed results in the five villages. They were effective in three villages: Tomoroco, Kaynakas and Talahuanca. The most positive outcome is that in these villages (on average) more than 80% of the families are currently involved in one way or another in SWC activities, and this was achieved without using incentive schemes or cash prizes. Most villagers also consider the executed SWC practices useful, and more than half of the farmers plan to replicate more measures in the near future. The three villages have in common that they all have solid foundations for sustainable development, which were laid before the SWC contests were conducted (in the first Phase of the logical strategy). This has triggered a renewed interest in alternatives to improve living conditions, including better soil management. The most negative outcomes are that in two of these villages (Kaynakas and Talahuanca) replication rates are currently very low, and in all villages CLs are no longer active as trainers. It seems that the dynamics of the process came to a halt after the project’s withdrawal. Some farmers cautiously experiment and replicate SWC practices, but most of them only maintain existing practices and wait for tangible results before investing in new ones. Similarly, the CLs find themselves in a vacuum; they are rarely asked for advice, and there is no common objective to keep the CLs’ groups active. The catalyst of the process, the project, is no longer there. The CLs were expected to fulfill this motivating role after the project’s withdrawal, but this has not happened.
On the other hand, in Tomoroco and Kaynakas the landscape has visibly changed due to the installed SWC practices, and internal regulations concerning controlled grazing are being complied with (Kessler 2007a). According to the respondents in the ex-post evaluation, SWC contests work; 80% of respondents are positive about the actual impact of the contests. Most importantly, for many farmers the experiences acquired during the contests with alternative techniques and practices have served as a basis for experimentation. Under the marginal conditions of poor farmers, adapting innovations is more important than adopting innovations (Van de Fliert and Braun 2002). The message that alternatives are available to improve productivity has come through; people are interested and have started to experiment with new techniques.
In this respect, it is interesting to observe the usage of other SWC practices that were not included in the contests, but are now practiced by interested farmer groups and CLs. Table 4 shows that several of these practices are currently in use by a considerable number of families, especially improved traditional practices like crop rotations (including leguminous crops) and mixed cropping systems. Hence, knowledge transfer from farmer to farmer does also occur spontaneously. If these SWC practices would have been included in the SWC contests, their adoption rate could have been much higher. Therefore, a first recommendation of this paper is to also conduct SWC contests in which practices are not previously defined. The contests described in this paper filled an important knowledge gap and provided farmers with basic information. However, the failure of earth bunds and the removal of other practices by several families prove that only executing contests with some predefined practices is not enough. Stimulating people’s creativity and having each individual family decide which management practices best fit their conditions is considered crucial in the MARENASS and SID projects (Van Immerzeel and De Zutter 2005). Local adaptations of existing practices will become available sooner if more space is left for farmers’ initiatives during the contests. Hence, the JGRC-type of contests can be maintained as a training tool, but other (more general) SWC contests should be organized as a follow-up activity.
Table 4Percentage of families using other SWC practices two years after project withdrawalSWC practicesExperimental villagesValidation villagesObservationsTomorocoKaynakasTalahuancaPatallajtaManure use361650Improved traditional practiceGreen manure200100New practiceCrop rotations60283015Improved traditional practiceStrip cropping2412150New practiceAgroforestry28642545New practiceMixed cropping68487065Improved traditional practiceComposting4020–10New practiceSource: Ex-post evaluation data in 2005
A second recommendation is to strengthen the role of local organizations in providing follow-up support to the SWC contests. The major weakness of the JGRC project was its incapacity to institutionalize the process: SWC activities continued at a high rate when the project fulfilled its catalyst function, but drastically decreased after the project’s withdrawal. Of course, municipalities were always involved in the activities, but their genuine participation was not achieved. Longer-lasting institutional support in organizing and facilitating more SWC contests would have strengthened the CLs’ role, and would have kept the dynamic process going. To some extent, farmers can respond to land degradation without external support, but they need continued provision of technical assistance and information in order to make progress (Paudel and Thapa 2001). Farmers have often lost the self-confidence and capacity to adapt and innovate (Reijntjes and others 1998); without external support they will continue to farm in the way they have in the past (Percy 2005). Given that SWC contests are a low-budget extension tool, they can be easily organized by municipalities and local NGOs, for example. However, despite the involvement of local leaders during the SWC contests, the active involvement of many extension workers is needed, especially when larger areas are to be covered in order to achieve a wide-scale impact. This can be a major limitation for local institutions; policies at the macro-level that enable the implementation of a farmer-based extension approach are therefore required. Earlier in this paper we already mentioned that in Bolivia profound changes at institutional level are required that give priority to extension. Only when such changes are made can SWC contests become an effective extension tool “to make haste slowly.”
In Bolivia, to date, governments have never committed themselves to extension; strategies were not clearly defined and the extension service constituted a large burden on the state budget (Bojanic 2001). Presently, many participatory approaches and tools are available (Chambers and others 1989) that have proven their effectiveness over the last decades. Based on case studies in Thailand and Laos, Connell (2000) concludes that there are several opportunities for the institutionalization of participatory approaches in mainstream extension, but that they all require significant political commitment. Particularly regarding environmental problems, different interest groups often pull in complementary and opposing directions (Röling and Pretty 1997). The challenge ahead for effective extension is to combine efforts. This has been done in Chile, where the government has contracted private technology companies to cater for the larger commercial farmers, and NGOs for small subsistence-oriented farmers. Rivera and Qamar (2003) for example propose a mixture of funding and service delivery modalities; governments could provide funding and NGOs could deliver the extension services. If the political willingness is present, this could also present an interesting opportunity in Bolivia.
Conclusions
The challenge we face in SWC is to quickly achieve widespread sustainable results, i.e., “to make haste slowly” (Savenije and Huijsman 1991). Based on the success of SWC contests elsewhere in the Andes (Van Immerzeel and De Zutter 2005), the JGRC project used this innovative tool in five rural Bolivian villages to put “making haste slowly” into practice. Mixed results were achieved with SWC contests between farmer groups. On the one hand, in villages where a solid foundation for sustainable development had already been laid, participation rates in the SWC contests were high. Most farmers were still involved in SWC activities even two years after project withdrawal, without receiving any incentive. In these villages large areas are currently protected with physical SWC practices and farmers have also started to experiment with other soil management practices. On the other hand, in the same villages the renewed system of collaboration focused on SWC was lost when the project withdrew, and Conservation Leaders did not continue with their training activities. Despite the visible widespread impact of the contests, sustainability is thus not yet assured. Farmers easily become disillusioned and unmotivated in the absence of a catalyst to keep the process of SWC contests and farmer-to-farmer training going. Moreover, tangible results such as higher soil productivity take a long time to appear. Farmers are opportunistic; in poor regions like the Bolivian mountain valleys, especially, they will grasp any opportunity to increase income.
Two recommendations were given in this paper to make SWC contests more effective in an extension strategy. First, in addition to the contests described here, other SWC contests should be organized in which practices are not predefined; this will stimulate peoples’ creativity in developing adaptations of existing practices. Second, commitment is required from local institutions to support SWC contests as an extension tool; only then can Conservation Leaders continue their activities, and can a widespread impact be achieved. Given the responsibility of Bolivian municipalities for rural development, they must be the first to become actively involved in extension. However, steering and support with adequate strategies from departmental and state institutions is indispensable; this will motivate municipalities to take natural resources conservation and rural development more seriously. Without such support, any attempt to spread SWC practices via participatory extension methodologies – such as farmer contests – will likely fail; no matter how logical and well-designed the strategy may be.

soil and water conservation
bolivian mountain valleys
farmer-to-farmer training
farmer-based extension
farmer contests
sustainable rural development