It was previously shown that nicotinic receptor activation induces detrusor muscle contractions via facilitating fast synaptic transmissions between preganglionic and postganglionic neurons (1, 2). In a canine bladder, it has been reported that smooth muscle contractility is preserved after decentralization indicative of functional muscarinic receptors (3). Here, utilizing ex vivo studies, we were interested in evaluating alterations in the expression of nicotinic receptors mediating bladder function after long-term decentralization in canines.

This study is an add-on investigation to a long-term project designed to develop novel surgical approaches for restoration of bladder, urethral and anal sphincter control to lower motoneuron lesioned pelvic organs.  Four female canines were decentralized by bilateral transection of all coccygeal and sacral (S) spinal roots, dorsal roots of lumbar (L)7 and hypogastric nerves then euthanized 11-21 months post-decentralization. Controls included sham-operated and unoperated animals. Mucosa-denuded smooth muscle strips were suspended in muscle baths and the maximal contractile responses to 120 mM potassium chloride (KCl) were measured. Then, strips were incubated with different antagonists for 20 minutes. Next, contractions to nicotinic receptor agonist epibatidine (10μM) were determined. Responses are expressed as percentages of KCl-induced contraction.

While the muscarinic receptor antagonist atropine (1μM) completely blocked epibatidine in muscle strips from both control and decentralized bladders, the sodium channel blocker tetrodotoxin (TTX, 1μM) inhibited epibatidine-induced contractions in strips from decentralized, but not control bladders. Competitive neuromuscular nicotinic receptor antagonists atracurium (5μM) or tubocurarine (0.1μM) inhibited epibatidine induced contractions in strips from control, but not decentralized bladders. The neuronal (α3β4) selective antagonist SR16584 (3-10μM) inhibited epibatidine contractions in both groups.

Atropine blockade of epibatidine contractions indicates that these nicotinic receptors induce release of acetylcholine that acts on muscarinic receptors to induce bladder muscle contraction. Because TTX does not block epibatidine contractions in strips from control bladders, this suggests that nicotinic receptors are likely located on nerve terminals and their antagonist pharmacology suggests that these consist of both α3β4 neuronal and (α1)2β1δε neuromuscular subtypes.  TTX blockade of contractions in the decentralized group suggests that long term decentralization causes nicotinic receptors to relocate from the nerve terminals to along the axons of the presynaptic neurons distant from the neuromuscular junction such that agonist induced depolarization triggers TTX sensitive action potentials that induce acetylcholine release at the neuromuscular junction.  The antagonist pharmacology of these axonal nicotinic receptors is consistent with neuronal (α3β4) nicotinic receptor subtypes.

It is possible that changes in the nicotinic receptor profile after long-term decentralization might suggest a physiological relevance of these nicotinic receptors in the bladder as a compensatory mechanism to recover bladder function and a potential target for drug treatment.

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Frara N, Giaddui D, Braverman AS, et al., 2021. Nerve transfer for restoration of lower motor neuron-lesioned bladder function. Part 2: Attenuation of purinergic bladder smooth muscle contractions. Am J Physiol Regul Integr Comp Physiol.