Urinary bladder function depends on autonomic innervation and a complex neural control system consisting of the brain and spinal cord. Its reliance on central nervous control distinguishes the bladder from other visceral structures such as those found in the cardiovascular and gastrointestinal systems. The efficient regulation by afferent and efferent activity in the urinary bladder is critical in its unusual task of both storage and intermittent voiding. And the appropriate postjunctional response to upstream neural regulation is imperative to mediate normal urinary voiding function. A major neurotransmitter influencing bladder contractile function is acetylcholine, which acts via M₂ and M₃ muscarinic receptors in smooth muscle. The M₃ receptor mediates direct contraction, whereas the M₂ receptor has been shown to enhance M₃-mediated function and to inhibit relaxation mediated by adenylate cyclase.

Diabetes is the most common cause of autonomic neuropathy in the developed world. Accordingly, visceral structures such as the urinary bladder are affected, where inadequate and aberrant control leads to dysfunction. In addition to neuropathy, diabetic bladders are also associated with distension. These abnormalities are reproduced in animals treated with streptozotocin (STZ), a pancreatic β-cell toxin. Through circumstantial evidence showing substantial enlargement in bladder from STZ-treated M₂ muscarinic receptor knockout (M ₂ KO) mouse, we speculated that excessive bladder distension might impose physical strain on surrounding nerves already under hyperglycemic stress. An upregulation of M₂ receptor expression and an increase in muscarinic receptor function in diabetic bladder has been reported as well as the enhanced functional profile of the M₂ receptor when the bladder becomes distended.

We hypothesize, therefore, that the M₂ receptor may have a protective role and may sustain function in the impaired diabetic bladder by opposing neuropathy secondary to excessive bladder distension. We postulate that M₂-mediated postjunctional compensatory mechanisms may also contribute to maintaining bladder function. We test these hypotheses in the following sections. In Chapter 2, we confirm that known postjunctional M ₂ receptor-mediated mechanisms in bladder can be replicated in an electrical field stimulation experiment, which acts as a reporter assay for M₂ receptor-mediated neurogenic bladder function. Chapter 3 investigates the differential postjunctional roles of M₂ and M₃ receptors in STZ-induced diabetes. Bladders are isolated from wild type and M₂ KO mice treated with vehicle or STZ. Contractions elicited to the bath-applied muscarinic agonist oxotremorine-M are measured in the absence and presence of the adenylate cyclase stimulator forskolin. We find that M₂-mediated responses sustain bladder contraction when M₃-mediated function is impaired in STZ-induced diabetes. And the ability of the M₂ receptor to inhibit the relaxant effects of forskolin is increased in this model. Chapter 4 shows that bladder lacking the M₂ receptor is pre-disposed to excessive enlargement during the course of STZ-induced diabetes. We hypothesize that an over-distended bladder phenotype may impinge surrounding nerves and, thereby, expedite the progression of diabetic neuropathy. Chapter 5 studies whether the contractile role of the M₂ receptor in gastric fundus is similar to that in bladder during the course of diabetes, since the fundus shares with the bladder the quality of being able to distend. The ileum does not share this quality to the same extent and so is not expected to exhibit a similar role for the M₂ receptor. Atrial contraction is also examined to determine muscarinic receptor involvement in another important system affected by diabetes. In Chapter 6, we discuss various possibilities, both theoretical and empirically explored by my research, by which the M ₂ receptor may mediate its protective role during the course of diabetes.