Neuronal regulation of myenteric interstitial cells of Cajal (ICC-MY) in the proximal colon

Abstract

Interstitial cells of Cajal (ICC) generate contractile patterns of colonic motility. We investigated innervation of ICC within the plane of the myenteric plexus (ICC-MY) in proximal colon using mice expressing GCaMP6f in ICC. ICC-MY generated localized Ca²⁺ transients that couple to activation of ANO1 channels, a Ca²⁺-activated Cl^- conductance. ICC are electrically coupled to SMCs, so activation or suppression of currents in ICC affects excitability of SMCs. ICC-MY displayed tonic inhibition, as the neurotoxin, TTX, increased the frequency of Ca²⁺ transients. Tonic inhibition was mimicked by a nitric oxide donor, NONOate, and by a guanylate cyclase agonist (Bay 58–2667). In contrast ODQ mimicked effects of TTX, increasing Ca²⁺ transients. Carbachol (CCh) increased Ca²⁺ transients in ICC-MY, and these effects were mediated by M3 muscarinic receptors. Neostigmine, also increased Ca²⁺ transients, suggesting there is tonic activation of enteric excitatory neurons in colonic muscles. Substance P and antagonists of NK1 and NK2 receptors had no effect on Ca²⁺ transients in ICC-MY. Electrical field stimulation (EFS), under conditions that emphasized excitatory neural responses, enhanced Ca²⁺ transients, and these effects were blocked by atropine or an M3 receptor antagonist (DAU 5884). EFS in the presence of atropine caused inhibition of Ca²⁺ via release of NO. Cessation of nitrergic stimulation resulted in a substantial increase in Ca²⁺ transients, known as post-stimulus excitation. In summary, ICC-MY, important for the generation of propulsive contractions in the colon, are innervated by excitatory (cholinergic) and inhibitory (nitrergic) motor neurons, and these inputs regulate the excitability of these cells.