Alterations of Colonic Neuromuscular Excitatory Cholinergic Pathway in a Rat Model of 6-Hydroxydopamine-Induced Parkinson's Disease

Introduction. Parkinson's disease (PD) is characterized by degeneration of nigrostriatal dopaminergic neurons. Patients with PD can develop gastrointestinal motor dysfunctions and alterations of their enteric nervous system, although the underlying mechanisms are poorly understood. The aim of the present study was to examine the patterns of colonic neuromuscular excitatory cholinergic pathway in a rat model of PD. Methods. PD was induced in male rats by intra-nigral injection of 6-hydroxydopamine (6-OHDA). Animals were sacrificed at 28 and 56 days after surgery. Colonic longitudinal muscle preparations were set up in organ baths with Krebs solution, and connected to isometric transducers to record contractions (g/g tissue) elicited by electrical stimulation (ES, 1-20 Hz). Cholinergic responses to ES were obtained in the presence of guanethidine (10 μM), Nω-nitro-L-arginine methylester (LNAME, 100 μM) and L-732,138 (NK1 receptor antagonist, 10 μM). Contractions elicited by carbachol (0.001-100 μM) in the presence of tetrodotoxin (1 μM) were also recorded. The concentrations of acetylcholine released into the incubation medium of colonic preparations were measured by fluorimetric assay, under resting conditions and 10 s after application of ES (10 Hz). The expression of choline acetyltransferase (ChAT) in the colonic neuromuscular layer was assessed by immunohistochemistry. Results. In control preparations, ES (10 Hz) evoked atropine-sensitive cholinergic contractions (49.6±5.3 g/g tissue), which were reduced in rats with PD at 28 and 56 days (31.3±4.6 and 37.7±2.3 g/g tissue, respectively). Contractions evoked by carbachol (1 μM) were enhanced in rats with PD both at 28 and 56 days (92.6±3.3 and 91.7±2.4 g/g tissue, respectively), as compared with controls (51.7±4.4 g/g tissue). Acetylcholine levels in the incubation medium of control tissues under resting conditions accounted for 11.8±0.7 μM/g, while they were 27.1±3.1 μM/g 10 s after ES application. In the medium of tissues from rats sacrificed 56 days after PD induction, acetylcholine levels did not vary vs controls under resting conditions, while they were reduced 10 s after ES application (-55% vs control). ChAT immunopositivity was detected in the myenteric ganglia of control tissues. In the colonic neuromuscular compartment of PD rats sacrificed both at 28 and 56 days, ChAT expression was significantly reduced, as compared with controls. Conclusion. Experimental PD is associated with an impairment of the colonic excitatory neurogenic cholinergic motility and an increase in the myogenic cholinergic contractile activity (likely mediated by a compensatory upregulation of muscarinic receptors). Such alterations appear to depend on a decrease in the density of myenteric cholinergic neurons, resulting in a reduction of acetylcholine release.