Sympathetic inhibition of ascending and descending interneurones during the peristaltic reflex in the isolated guinea-pig distal colon.

We investigated the effects of sympathetic nerve stimulation within ascending and descending reflex pathways underlying the peristaltic reflex in the guinea-pig distal colon. A three-chambered partitioned bath was used to divide a segment of distal colon into stimulation, recording and intermediate regions. The effects of lumbar colonic nerves (LCN) could be localized to the intermediate region by surgical lesions of the mesentery and by application of guanethidine (3 μm) to the stimulation and recording chambers. Brush stroking the mucosa in the anal and oral stimulation chambers elicited a synchronous contraction of the longitudinal muscle (LM) and circular muscle (CM) oral to, and transient relaxation of the LM and CM anal to, the stimulus, respectively. After Nω-nitro-L-arginine (L-NA; 100 μm) in the oral and intermediate chambers, mucosal stimulation in the oral chamber elicited a prolonged descending inhibitory and excitatory complex in both the LM and CM in the anal recording chamber. This was blocked by hexamethonium (300 μm), which did not affect the transient relaxation response recorded in control conditions. Stimulation of the LCN (1200 pulses, 20 Hz), delivered to the intermediate region, abolished the oral contraction and the L-NA-induced anal complex in both the LM and CM, but was without effect on the transient hexamethonium-resistant anal relaxation. These effects of LCN stimulation were reversed by phentolamine (3 μm) or yohimbine (100 nM), but not propranolol (10 μm), when added to the intermediate chamber. LCN stimuli (2-20 Hz, 600 μs pulses) directed to the recording chamber elicited synchronous relaxations in the LM and CM that were unaffected by hexamethonium (300 μm), but were reduced by yohimbine and usually blocked by the further addition of propranolol (10 μm). In conclusion, sympathetic nerve stimulation inhibits orally and anally projecting cholinergic interneurones underlying the peristaltic reflex in the distal colon. In addition, the LM and CM relax synchronously following release of sympathetic neurotransmitter, over a range of stimulus frequencies. It is generally believed that stimulation of the sympathetic nervous system generates an inhibitory action on the movements of the large intestine (Bayliss & Starling, 1900; Garry & Gillespie, 1955; Gillespie, 1962; Bennett et al. 1966; Mackenna & McKirdy, 1972; Furness & Costa, 1987; Szurszewski & Miller, 1994). There is considerable evidence that this inhibition is mediated largely via an indirect action on the enteric nervous system, rather than directly on the smooth muscle. This has been supported, in many species, by immunohistochemical studies that have revealed nerve terminals with intense immunoreactivity for tyrosine hydroxylase surrounding neurones within the myenteric plexus, and a relatively sparser innervation in the smooth muscle (Norberg, 1964; Furness, 1970; Furness & Costa, 1973; Gabella, 1979; Burnstock & Wong, 1981; Spencer et al. 1998a). In addition, during direct intracellular recordings from myenteric neurones it has been shown that stimulation of sympathetic fibres inhibits spontaneous and evoked fast synaptic transmission within the myenteric plexus (Hirst & McKirdy, 1974), and there is considerable evidence that this is likely to occur via presynaptic inhibition of the release of acetylcholine (Hirst & McKirdy, 1974; Tack & Wood, 1992). However, it is unclear which myenteric neurones receive sympathetic inputs in the colon and, moreover, the role of this innervation during the peristaltic reflex in the large bowel is unclear. In addition to a lack of knowledge of the classes of enteric neurone under sympathetic neuronal control, the effects of sympathetic stimulation on the relative movements of the circular muscle (CM) and longitudinal muscle (LM) layers have received little attention. Since the late 1960s, there has been some controversy associated with the relative movements of the CM and LM layers in the gastrointestinal tract (see the introductions in McKirdy, 1972 and Smith & Robertson, 1998). Kottegoda (1969, 1970) put forward the once generally accepted idea of reciprocal innervation, i.e. when the CM layer contracts, the LM relaxes during peristalsis and vice versa. However, he appears to have overlooked any possibility of passive mechanical interactions that may occur between the movements of the LM and CM, which can lead to false interpretations of their relative movements (see Gregory & Bentley, 1968; Wood & Perkins, 1970; McKirdy, 1972). When dissection techniques are employed to prevent such mechanical interactions between the muscle layers, they are found to contract and relax synchronously in response to a peristaltic wave (Smith & Robertson, 1998) and reflex stimulation of the mucosa (colon: Smith & McCarron, 1998; ileum: Spencer et al. 1999). However, in response to stimulation of extrinsic nerves, the relative movements of the LM and CM layers is less clear. Some investigators observed that both muscle layers of the large intestine relaxed in response to sympathetic nerve stimulation (canine large intestine: Bayliss & Starling, 1900; rabbit large intestine: Garry & Gillespie, 1955; Mackenna & McKirdy, 1972) but contracted in response to pelvic nerve stimulation (McKirdy, 1972). In some studies, however, it has been reported that sympathetic nerve stimulation elicits contractions in the CM layer (Lee, 1960; Venkova & Krier, 1993; Luckensmeyer & Keast, 1998) but relaxation of the LM (Lee, 1960; Luckensmeyer & Keast, 1998) of the proximal colon. In these latter studies, it is unclear whether the two muscle layers were sufficiently mechanically isolated from each other, so that the actual movements of the circular and longitudinal muscles could be studied unambiguously following extrinsic nerve stimulation. In the current study, we have used our previously described dissection techniques (Smith & Robertson, 1998; Smith & McCarron, 1998; Spencer et al. 1999) to unambiguously investigate the effects of sympathetic nerve stimulation on the relative movements of the LM and CM layers of the colon. Furthermore, we have used the partitioned organ bath technique to localize the effects of sympathetic nerve stimulation to particular neural elements within ascending and descending reflex pathways.