Gastrointestinal function is frequently abnormal in patients with poorly controlled diabetes. Clinical investigations indicate that most of these abnormalities can be attributed to defective vagal afferent functioning. Our preliminary studies indicate that nodose ganglia (NG) neurons in streptozotocin induced diabetic rats (STZ- D) display hyperpolarization leading to decreased excitability. This may contribute to abnormal vagal function in the diabetic state. We hypothesize that chronic hyperglycemia activates "background" TRESK potassium channels, leading to hyperpolarization of the NG and decreased excitability. This is a two-step process. Initially increase in intracellular calcium in diabetic neurons activates calcineurin. Calcineurin binds to a NFAT- like docking site on the TRESK protein and causes dephosphorylation of serine 276, resulting in activation of the channel and leading to hyperpolarization. Over time upregulation of the TRESK protein occurs resulting in not only increased frequency of the opening of TRESK channel but an increase number of TRESK K+ channels. To test this hypothesis we have 3 specific aims.
Aim 1 is to demonstrate that hyperglycemia in STZ- D modifies basic electrophysiological properties of NG neurons. Patch clamp recordings will be performed to characterize the excitability of NG ganglia neurons from control and STZ-D rats. Physiological implications of these abnormalities will be evaluated by in vivo electrophysiological recording of NG in diabetic rats and study its responsiveness to CCK, leptin and secretin stimulation.
Aim 2 examines whether hyperpolarization of NG neurons in the chronic diabetic state is mediated by activation of TRESK channels. The presence of specific TESK potassium channels in NG will be identified using electrophysiological studies as well as western blot and RT-PCR methods. The participation of the TRESK channel will be demonstrated by the use of a virus based system for delivery of siRNA to silence the expression of the TRESK channel in NG. To evaluate the functional importance of TRESK in the mediation of hyperpolarization of NG in diabetes, we will examine reversibility of the electrophysiological and GI abnormalities following silencing TRESK channel expression in vivo through electroporation of the NG with TRESK siRNA.
Aim 3 investigates the signal transduction cascades that mediate the membrane modifications of NG neurons in diabetes. Patch clamp recordings and intracellular calcium imaging studies will be performed with messenger specific activators or inhibitors to determine the role of specific intracellular cascade elements on NG excitability in diabetic animals. The demonstration of desphosphorylation of serine 276 in diabetes will involve the use of phosphoproteome method. Understanding the cellular and molecular mechanism responsible for abnormal functioning of the NG in the diabetic state will provide important therapeutic targets for the management of abnormal GI function in chronic diabetes.

Public Health Relevance

Gastrointestinal functions are frequently abnormal in patients with poorly controlled diabetes, including abnormal stomach motility, diminished pancreatic and stomach secretions and abnormal eating behavior. Many of these functions are controlled through a vagus nervous system which is frequently abnormal in diabetic patients. The purpose of this study is to understand the molecular and cellular mechanisms responsible for the abnormalities of this group of sensory neurons. This could provide important therapeutic targets and lead to improved management of diabetic patients with GI complications.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK084039-03
Application #
8257173
Study Section
Clinical, Integrative and Molecular Gastroenterology Study Section (CIMG)
Program Officer
Carrington, Jill L
Project Start
2010-05-01
Project End
2015-04-30
Budget Start
2012-05-01
Budget End
2013-04-30
Support Year
3
Fiscal Year
2012
Total Cost
$319,436
Indirect Cost
$114,011
Name
University of Michigan Ann Arbor
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
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