bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effects on K+ currents in rat cerebellar granule neurones of a membrane‐permeable analogue of the calcium chelator BAPTA
Authors: Watkins Christopher S. , Mathie Alistair 
Published: July 2012

. Whole cell recordings of voltage‐activated K+ currents were made with the amphotericin B perforated patch technique from cerebellar granule (CG) neurones of 6–8 days rats that had been in culture for 1 to 16 days. By use of appropriate voltage protocols, the effects of the membrane‐permeant form of BAPTA, 1,2‐bis‐(2‐amino‐phenoxy)ethane‐N,N,N′,N′‐tetraacetic acid acetoxymethyl ester (BAPTA‐AM), on the transient A current (IKA), the delayed rectifier current (IKv) and a standing outward current (IKso) were investigated.
. Bath application of 25 μm BAPTA‐AM inhibited both IKv and IKso in cultured neurones, but did not seem to affect IKA. Neither 25 μm BAPTA (free acid) nor 25 μm ethylenediaminetetraacetic acid acetoxymethyl ester (EDTA‐AM) had any significant effect on the magnitude of IKso. Similarly in short‐term (1–2 days) cultured CG neurones IKv, but not IKA, was inhibited by 25 μm BAPTA‐AM.
. BAPTA‐AM (2.5 μm) reduced IKv in short‐term culture CG neurones, with further inhibition being seen when the perfusate was changed to one containing 25 μm BAPTA‐AM.
. Tetraethylammonium ions (TEA) (10 mM) reversibly inhibited IKv in these cells with a similar rate of block of IKv to that induced by 25 μm BAPTA‐AM.
. The degree of inhibition of IKv by 25 μm BAPTA‐AM was both time‐ and voltage‐dependent, in contrast to the inhibition of this current by TEA.
. These data indicate that BAPTA‐AM reduces K+ currents in cerebellar granule neurones and that this inhibition cannot be explained in terms of intracellular Ca2+ chelation, but is a direct effect on the underlying channels.
Direct chronotropic effects of atrial and C‐type natriuretic peptides in anaesthetized dogs
Authors: Beaulieu Pierre , Cardinal René , De Léan André , Lambert Chantal 
Published: July 2012

. The chronotropic effects of atrial natriuretic peptide (ANP) and C‐type natriuretic peptide (CNP) were investigated using injections (50 μg in 1 ml of Tyrode solution as bolus over 1 min) directly into the sinus node artery of 21 anaesthetized and vagotomized dogs which had been pretreated with a β‐adrenoceptor antagonist. The injections were also repeated following: (a) α‐adrenoceptor antagonism (prazosin) and muscarinic receptor antagonism (atropine); (b) inhibition of prostaglandin synthesis (indomethacin); (c) angiotensin II AT1 receptor antagonism (losartan); (d) histamine H1 (mepyramine) and H2 (cimetidine) receptor antagonism.
. The results obtained indicate that ANP had no significant effect on the basal sinus rate, whereas CNP produced a slight but significant increase of 12 ± 2 beats min−1. The effect of CNP was long‐lasting (return to pre‐injection levels after maximum effect in 17 ± 3 min) and was not influenced by the various antagonists mentioned above.
. During in vitro experiments on spontaneously beating right atria isolated from 6 dogs, the injection of CNP (50 μg in 1 ml of Tyrode solution) into the sinus node artery produced an increase in atrial rate of 14 ± 1 beats min−1.
. The results of this work indicate that CNP exerts a significant and prolonged positive chronotropic effect both in vivo and in vitro. Other studies are required to elucidate the mechanism of action of CNP on the heart conduction system, to ascertain the presence of natriuretic peptide receptor B in the region of the sinoatrial node and to determine the role of CNP in the control of heart rate.
Modulation by calcineurin of 5‐HT3receptor function in NG108‐15 neuroblastoma x glioma cells
Authors: Boddeke H.W.G.M. , Meigel I. , Boeijinga P. , Arbuckle J. , Docherty R.J. 
Published: July 2012

. We have investigated the mechanism of regulation of 5‐HT3receptor channel sensitivity in voltage‐clamped (−80 mV) NG108‐15 neuroblastoma cells.. The 5‐HT‐induced inward current activated rapidly. The fast onset was followed by a biphasic decay which was characterized by two time constants, τ1(1.1 ± 0.21s) and τ2(8.9 ± 1.6s), respectively. Brief applications of 5‐HT, applied at 2 min intervals, induced a decrease in the amplitude of the 5‐HT3receptor‐mediated peak inward currents.. Buffering of intracellular calcium with the calcium chelator BAPTA (10 mM) instead of EGTA (10 mM) attenuated the 5‐HT‐induced loss of responsiveness of 5‐HT3receptors. Omission of calcium from the extracellular medium yielded a similar attenuation of loss of responsiveness.. Inclusion of the protein kinase inhibitor, staurosporine (1 μm) or of okadaic acid (1 μm), an inhibitor of protein phosphatases 1 and 2A, in the intracellular buffer solution did not affect 5‐HT3receptor sensitivity.. Injection of cyclosporin A‐cyclophilin A complex (20 nM), which potently inhibits calcineurin, did not affect the time constants of the biphasic decay of the 5‐HT response τ1(1.4 ± 0.28s) and τ2(11.3 ± 1.7s). The complex, however, prevented the loss of 5‐HT3receptor responsiveness upon repeated application of 5‐HT. A similar, but weaker effect was observed after intracellular application of the autoinhibitory peptide domain of calcineurin (1 μm).. The recovery of desensitized 5‐HT3receptors upon a second application of 5‐HT (1 μm) showed a half‐life time (τ1/2) of 2.6 ± 0.12 min in control cells which was reduced to 1.6 ± 0.09 min in cells treated with cyclosporin A‐cyclophilin A (20 nM) complex.. We conclude that calcineurin does not affect the fast decay of the 5‐HT3receptor response but may be involved in a slower process which regulates channel activity.
Evidence for P‐glycoprotein‐modulated penetration of morphine‐6‐glucuronide into brain capillary endothelium
Authors: Huwyler Jörg , Drewe Jürgen , Klusemann Caterina , Fricker Gert 
Published: July 2012

Morphine‐6‐glucuronide is one of the major metabolites of morphine. The potent analgesic action of this compound together with its potential lower apparent toxicity in man, when compared with morphine, indicated its clinical importance.
Primary cultures of porcine brain capillary endothelial cells were used to study brain penetration of morphine‐6‐glucuronide. Biochemical characterization of the cell cultures revealed a marked enrichment in enzymatic activity of alkaline phosphatase (56 fold) and angiotensin converting enzyme (230 fold) as compared to whole brain tissue. By immunostaining the presence of vimentin, factor VIII, the tight junction associated protein ZO‐1, and P‐glycoprotein was shown. Functional characterization revealed that the carrier system responsible for transport of neutral amino acids was intact.
Uptake and transport of morphine‐6‐glucuronide was marginal and in the range of the extracellular marker sucrose. However, uptake of morphine‐6‐glucuronide was enhanced significantly (P<0.0001) in presence of the inhibitors of P‐glycoprotein, verapamil or vincristine. The finding that morphine‐6‐glucuronide may serve as a substrate for P‐glycoprotein was confirmed in multidrug‐resistant P388 tumour cells.
We conclude that penetration of the blood‐brain barrier by morphine‐6‐glucuronide may depend on the expression of the product of the multidrug‐resistance (MDR) gene in brain capillary endothelial cells.
Effects of RP 73401, a novel, potent and selective phosphodiesterase type 4 inhibitor, on contractility of human, isolated bronchial muscle
Authors: Naline E. , Qian Y. , Advenier C. , Raeburn D. , Karlsson J.‐A. 
Published: July 2012

The aim of this study was to investigate the smooth muscle relaxant effects of the novel, selective phosphodiesterase (PDE) type 4 inhibitor, RP 73401 in comparison with the classical PDE 4 inhibitor, rolipram, the non‐selective PDE inhibitor, theophylline and the β‐adrenoceptor agonist, isoprenaline on the human, isolated bronchus.
At resting tone, the rank order of potency (pD2) for the relaxants was RP 73401 ≥ rolipram ≥ isoprenaline > > theophylline. In terms of maximum relaxation produced (Emax) the PDE 4‐selective inhibitors were similar, but the maximal effects (70–75% of theophylline, 3 mM) were lower than that observed with isoprenaline (98% of theophylline, 3 mM) or theophylline itself (100%).
On the human isolated bronchus pre‐contracted with acetylcholine (ACh, 0.1 or 1.0 mM), the rank order of potency remained the same. The maximal responses to RP 73401 and rolipram were however markedly reduced (Emax 39.9–46.6%) compared with isoprenaline (Emax 79–85%).
In tissues pre‐contracted with ACh (0.1 mM), RP 73401 and rolipram (10−9‐10−7 m) significantly and concentration‐dependently increased tissue sensitivity to isoprenaline. RP 73401 and rolipram were similar in potency. Both selective PDE 4 inhibitors also significantly increased the maximal relaxant effects of isoprenaline. These effects were not observed with the PDE 3 inhibitor, siguazodan.
In terms of retention by tissues (an index of duration of action), the onset of action of RP 73401 (2.11 ± 0.53 min) and rolipram (1.70 ± 0.45 min) was significantly slower than that of isoprenaline (0.33 ± 0.06 min) or theophylline (1.17 ± 0.25 min). The retention of RP 73401 (89.0 ± 21.9 min) on the human isolated bronchial tissues after washing was however dramatically longer than that of rolipram (18.3 ± 4.5 min), theophylline (3.43 ± 0.58 min) or isoprenaline (2.81 ± 0.31 min).
These data indicate that RP 73401 is a potent and long‐acting relaxant of human bronchial muscle in vitro. RP 73401 is more potent than the classical PDE 4‐selective inhibitor rolipram and the non‐selective PDE inhibitor theophylline and is retained in bronchial tissue for a much longer period of time.
Effect of intrarenally infused parathyroid hormone‐related protein on renal blood flow and glomerular filtration rate in the anaesthetized rat
Authors: Massfelder Thierry , Parekh Niranjan , Endlich Karlhans , Saussine Christian , Steinhausen Michael , Helwig Jean‐Jacques 
Published: July 2012

Parathyroid hormone‐related protein (PTHrP) is expressed in the kidney and acts on vascular PTH/PTHrP receptors to vasodilate the isolated kidney and to stimulate renin release. However, effects of PTHrP on renal blood flow (RBF) and glomerular filtration rate (GFR) in vivo have not been assessed in the absence of its cardiac, peripheral and central effects. We investigated the renal effects of PTH and PTHrP infused into the left renal artery of anaesthetized rats.
Intrarenal infusions, adjusted to generate increasing concentrations of human PTHrP(1–34) and rat PTH(1–34) in renal plasma (2 × 10−11 to 6 × 10−9 m) produced a comparable dose‐dependent increase in RBF. The rise was 4% at the lowest and 34% at the highest concentrations of peptides. Up to a concentration of 2 × 10−9 m, mean arterial pressure (MAP) and heart rate were not affected, but at 6 × 10−9 m, intrarenally infused peptides reached the peripheral circulation, and caused a fall in MAP within a few minutes. While MAP returned to basal value after the last peptide infusion, RBF remained more than 10% above control for at least 30 min.
Two competitive PTH/PTHrP receptor antagonists, [Nle8,18, Tyr34]‐bPTH(3–34)amide and [Leu11, D‐Trp12]‐hPTHrP(7–34)amide (2 × 10−8 m) were devoid of agonist activity, but markedly antagonized the dose‐dependent increase in RBF elicited by PTHrP.
GFR and urine flow were measured in left PTHrP‐infused experimental kidney and right control kidney. Renal PTHrP concentration of 10−10 m elevated left RBF by 10%, and GFR by 20% without significantly increasing filtration fraction, and increased urine flow by 57%. In the right control kidney GFR and diuresis did not change.
The results indicate that PTHrP has similar renal haemodynamic effects as PTH and increases RBF, GFR and diuresis in anaesthetized rats.
Impaired activation of adenylyl cyclase in lung of the Basenjigreyhound model of airway hyperresponsiveness: decreased numbers of high affinity β‐adrenoceptors
Authors: Emala C.W. , Aryana A. , Hirshman C.A. 
Published: July 2012

To evaluate mechanisms involved in the impaired β‐adrenoceptor stimulation of adenylyl cyclase in tissues from the Basenji‐greyhound (BG) dog model of airway hyperresponsiveness, we compared agonist and antagonist binding affinity of β‐adrenoceptors, β‐adrenoceptor subtypes, percentage of β‐adrenoceptors sequestered, and coupling of the β‐adrenoceptor to Gsα in lung membranes from BG and control mongrel dogs. We found that lung membranes from the BG dog had higher total numbers of β‐adrenoceptors with a greater percentage of receptors of the β subtype as compared to mongrel lung membranes.
Agonist and antagonist binding affinity and the percentage of β‐adrenoceptors sequestered were not different in BG and mongrel dog lung membranes. However, the percentage of β‐adrenoceptors in the high affinity state for agonist was decreased in BG lung membranes suggesting an uncoupling of the receptor from Gsα.
Impaired coupling between the β‐adrenoceptor and G protein documented by the decreased numbers of β‐adrenoceptors in the high affinity state in BG lung membranes, is a plausible explanation for the reduced stimulation of adenylyl cyclase and the resultant reduction in airway smooth muscle relaxation in this model.
Tachykininergic synaptic transmission in the coeliac ganglion of the guinea‐pig
Authors: Zhao F.‐Y. , Saito K. , Yoshioka K. , Guo J.‐Z. , Murakoshi T. , Konishi S. , Otsuka M. 
Published: July 2012

The responses of coeliac ganglion neurones of the guinea‐pig to electrical stimulation of the mesenteric nerves and applications of tachykinin receptor agonists were investigated by use of intracellular recording techniques.
Ganglion neurones were classified into three groups based on firing patterns in response to a depolarizing current pulse: phasic (38% of the population), tonic (39%) and atypical (23%). In the majority of phasic neurones (91%) a long after‐hyperpolarization (LAH) lasting 5–8 s followed action potentials induced by a train of depolarizing current pulses. In contrast, LAH was rarely observed in tonic neurones (5%).
In most of tonic neurones (90%) slow excitatory post‐synaptic potentials (e.p.s.ps) lasting 3–10 min were evoked by repetitive electrical stimulation of the mesenteric nerves. Prolonged depolarizations were also evoked in most tonic neurones by applications of substance P (SP), neurokinin A (NKA) or senktide, a tachykinin NK3 receptor agonist.
In most of phasic neurones (73%), mesenteric nerve stimulation did not induce an obvious depolarization but induced a prolonged inhibition of LAH lasting 3–10 min. Bath‐applied tachykinin receptor agonists similarly induced an inhibition of LAH without causing depolarization in most of the phasic neurones.
GR71251 (5 μm), a tachykinin NK1 receptor antagonist, partially depressed the nerve‐evoked slow e.p.s.ps in tonic neurones and the nerve‐evoked LAH inhibition in phasic neurones.
Capsaicin (0.1–5 μm) induced a prolonged depolarization in tonic neurones and an inhibition of LAH in phasic neurones.
A mixture of peptidase inhibitors potentiated the depolarization and the LAH inhibition evoked by nerve stimulation, SP and NKA, but not those evoked by senktide.
It is concluded that tonic neurones respond to repetitive mesenteric nerve stimulation preferentially with slow e.p.s.ps and that phasic neurones respond preferentially with LAH inhibition. The present study further suggests that SP and NKA, released from axon collaterals of primary afferent neurones, produce slow e.p.s.ps in tonic neurones and the LAH inhibition in phasic neurones via NK1 receptors.
Potentiation of chloride responses to glycine by three 5‐HT3 antagonists in rat spinal neurones
Authors: Chesnoy‐Marchais D. 
Published: July 2012

Modulations of Cl− responses to glycine by 5‐hydroxytryptamine ligands were studied in cultured spinal neurones, by the whole‐cell recording technique.
Three 5‐HT3 antagonists were found to potentiate reversibly responses to low concentrations of glycine. Potentiations were induced by micromolar concentrations of LY‐278,584 (1–10 μm) and by concentrations of MDL‐72222 or ICS‐205,930 between 10 nM and 1 μm.
Potentiations were observed over the whole voltage range without any change in the reversal potential of the glycine responses and without affecting the resting conductance.
The degree of potentiation was variable among cells. It increased with the concentration of the modulator, but only up to 100 nM for MDL‐72222 and ICS‐205,930.
The potentiation appeared to result from an increase in the affinity for glycine of glycine receptors.
Neither the blockade of glycine uptake by Na+ removal, nor the excision of membrane patches prevented the potentiation.
At high concentrations (10 μm), both MDL‐72222 and ICS‐205,930 had, in contrast, a blocking effect on glycine responses.
Potentiation by LY‐278,584 and a dose‐dependent modulation by MDL‐72222 were also observed for taurine responses.
The effects on glycine responses of various ligands of 5‐HT3 receptors (including agonists) are discussed. The ability of LY‐278,584, MDL‐72222 and ICS‐205,930 to potentiate glycine responses appears to be independent of their known 5‐HT3 receptor antagonist properties. It would be interesting to look for chemically related drugs that would be specific potentiators of glycine responses.
Facilitation of imipramine efflux from the brain by systemic specific antibodies
Authors: Ragusi Corinne , Boschi Gabrielle , Rips Richard , Scherrmann Jean‐Michel 
Published: July 2012

This study investigated the capacity of circulating anti‐tricyclic antidepressant (TCA) IgG to increase the efflux of imipramine (Imip) from the rat brain.
A tracer amount of [3H]‐Imip (40 pmol) was injected into the cerebral lateral ventricle and its efflux was determined in control rats and in rats given anti‐TCA antibody. The monoclonal anti‐TCA IgG1 was injected i.v. 48 h before Imip at 4 IgG:Imip molar ratios (10, 100, 1000 and 10000). The [3H]‐Imip in arterial and venous plasma was measured for up to 60 min, and in the brain and peripheral organs (heart, liver, lung, kidney) 5 and 60 min after Imip injection.
The arterial plasma concentration of Imip in control rats was significantly higher (26.7 ± 2.1 pM) than the venous one (17.7 ± 2.0 pM) at 5 min, indicating that Imip released from brain becomes distributed in peripheral tissues. These concentrations were not significantly different at 60 min suggesting that Imip was, at this time, redistributing from extravascular tissues to the blood. In rats given anti‐TCA IgG, any Imip leaving the brain was immediately bound by the circulating antibody at 5 min. This greatly reduced the Imip in the heart (63.9%) and lung (61.3%) at the highest IgG:Imip ratio. The brain Imip was markedly lower at 60 min (31.5% with an IgG:Imip ratio of 1000 and 57.5% at a ratio of 10000). The two lowest IgG:Imip ratios had less effect on the plasma Imip because of the relative low affinity of the anti‐TCA IgG (3.8 × 107 M−1).
These data indicate that the anti‐TCA IgG facilitated the efflux of Imip from the brain, even though these antibodies cannot cross the blood‐brain barrier. This may be an efficient system for increasing drug organ clearance, as more than half the Imip in the brain was actively removed by the antibody in 1 h.
Further evidence for the presence of cannabinoid CB1 receptors in guinea‐pig small intestine
Authors: Pertwee Roger G. , Fernando Susanthi R. , Nash Joanne E. , Coutts Angela A. 
Published: July 2012

CP 50,556, CP 55,940, nabilone, CP 56,667, Δ9‐tetrahydrocannabinol (THC) and cannabinol each inhibited electrically‐evoked contractions of the myenteric plexus‐longitudinal muscle preparation of guinea‐pig small intestine in a concentration‐related manner. The IC50 values of these cannabinoids, respectively 3.45, 3.46, 30.61, 162.94, 214.63, and 3913.5 nM, correlate well with previously obtained potency values for displacement of [3H]‐CP 55,940 from cannabinoid binding sites.
Electrically‐evoked contractions of the myenteric plexus‐longitudinal muscle preparation were also inhibited by AM 630 (6‐iodo‐pravadoline) and by WIN 55,212‐2 (IC50 = 1923.0 and 5.54 nM, respectively). The present finding that AM 630 is an agonist, contrasts with a previous observation that it behaves as a cannabinoid receptor antagonist in the mouse isolated vas deferens.
SR141716A produced dose‐related parallel rightward shifts in the log concentration‐response curves of CP 55,940, WIN 55,212‐2, THC and AM 630 for inhibition of electrically‐evoked contractions of the myenteric plexus‐longitudinal muscle preparation. SR141716A (1 μm) did not reverse the inhibitory effects of normorphine and clonidine on electrically‐evoked contractions or potentiate the contractile response to acetylcholine.
Doses of naloxone and yohimbine that reversed the inhibitory effects of normorphine or clonidine on electrically‐evoked contractions of the myenteric plexus‐longitudinal muscle preparation did not affect the inhibitory response to WIN 55,212‐2.
Electrically‐evoked release of acetylcholine from strips of myenteric plexus‐longitudinal muscle was decreased by 200 nM CP 55,940 and this inhibitory effect was almost completely reversed by 1 μm SR141716A. Acetylcholine‐induced contractions were not affected by 200 nM CP 55,940.
These results support the hypothesis that guinea‐pig small intestine contains prejunctional cannabinoid CB1 receptors through which cannabinoids act to inhibit electrically‐evoked contractions by reducing release of the contractile transmitter, acetylcholine.
THC was found to be more susceptible to antagonism by SR141716A than CP 55,940 or AM 630, raising the possibility that guinea‐pig small intestine contains more than one type of cannabinoid receptor.
At concentrations of 10 nM and above, SR141716A produced small but significant increases in the amplitude of electrically‐evoked contractions of the myenteric plexus‐longitudinal muscle preparation suggesting that this tissue may release an endogenous cannabinoid receptor agonist or that some cannabinoid receptors in this tissue are precoupled and that SR141716A can reduce the number of receptors in this state.