Type II diabetes: phenotype of the diabetic bladder from proteins to organ
35-50% of Americans with Type II diabetes experience diabetic bladder dysfunction (DBD) significantly reducing their quality of life. Our understanding of DBD has been hampered by the lack of an animal model that closely mirrors the human situation. The goal of this proposal is to develop a mechanistic phenotype of a novel model of Type II diabetes, the high fat diet (HFD) and low dose streptozotocin (STZ) rat. This model has superior construct validity as it utilizes compromised beta cells that are stressed by a HFD. The hypothesis to be tested is that the HFD/STZ rat can be used as an appropriate, non-genetic based animal model to study the consequences of Type II diabetes on bladder regulation and function. This hypothesis will be tested by the completion of three specific aims. Aim 1 will determine the optimal time for the study of pre-diabetic, compenated (defined as hyperactive bladder and hypercontractile smooth muscle), and decompensated (defined as atonic bladder and hypocontractile bladder) states of DBD using urodynamics and concious cystometry. Body weights, blood glucose, and plasma insulin will also be measured. Aim 2 will phenotype intact strips of bladder smooth muscle from the three states of DBD. Concentration-response curves to carbachol, bethanechol, ATP, and KCl and frequency-response curves to electrical field stimulation will be generated. Basal, peak, and steady state levels of myosin phosphatase, CPI-17, mitogen-activated protein kinase, and myosin light chain (MLC) phosphorylation will be quantified in response to maximal activation in strips from the three DBD states. Aim 3 will phenotype alpha-toxin permeabilized fibers in terms of the calcium dependence of force and MLC phosphorylation in response to calcium alone or in the presence of carbachol and GTP or GTPgammaS. The calcium dependence of force and MLC phosphorylation will also be determined using Triton X-100 detergent skinned fibers. This will provide information on the calcium sensitization pathways and the calcium dependence of the contractile proteins. Completion of these aims will provide a detailed mechanistic phenotype and provide the basis for future studies to determine the effect of a low fat diet and the development of novel therapeutic treatments.

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