Qun-Yong Zhou - US grants

DESCRIPTION (Adapted from applicant's abstract): The long-term objective of this project is to further our understanding of the physiological role of dopamine D1 and D2 receptors in normal CNS function and in the functional pathologies associated with schizophrenia and other mental health disorders. Evidence put forth by a number of investigators suggests that the overactivation of dopaminergic synaptic transmission may be important in the pathophysiology of schizophrenia. The precise relation of dopamine overactivation to schizophrenia, however, remaines vague. The present series of studies seeks to bridge this gap by investigating the consequences of overactivated dopamine systems on brain development and function by genetic manipulation of dopamine receptors, and by identifying the consequences of expression of mutant dopamine receptors in transgenic animals. In brief, these studies seek (a) to create constitutively active mutants of dopamine D1 and D2 receptors and to identify their pharmacological and biochemical characteristics, (b) to introduce selected constitutively active D1 and D2 receptor mutants into transgenic mice, (c) to characterize developmental abnormalities in animals with overactivated dopamine systems, and (d) to identify behavioral abnormalities in mice with constitutively active dopamine receptors. The studies will enhance our understanding of D1 and D2 receptors at the molecular level, lead to a better understanding of the action mechanisms for dopamine receptors, and may lead to the development of improved pharmacotherapies for dopamine system malfunctions. These studies will also lead to a better understanding of the developmental and functional consequences of dopamine overactivation and should provide us with some fundamental information pertaining to the relation of dopamine neurotransmission malfunctions with the development of the behavioral malfunctions underlying complex mental health disorders.

The mesolimbic dopamine system mediates the reinforcing effects of nicotine and other drugs of abuse. Thus the genes affecting the functionality of the mesolimbic dopamine system can be contributing factors to predisposition to nicotine abuse. These genes include dopamine receptors, signaling molecules of cAMP pathway such as adenylate cyclase, and others. Evidence put forth by a number of investigators suggests that the difference in the different biochemical profile of cAMP pathway in the mesolimbic dopamine system may result in differential predilection to abused drugs. The precise relation of up-regulated cAMP signaling pathway~y and nicotine predilection, however, remains vague. The present study seeks to bridge this gap by investigating the consequence of up-regulated cAMP pathway in transgenic mice, caused by a subtle substitution mutation in the dopamine D2 receptor, on nicotine responsiveness. Particularly, these studies seek to produce a strain of transgenic mice that express the mutant D2 receptor and to investigate whether the mutant D2 receptor will result in up-regulation of cAMP pathway in transgenic mice and therefore increased vulnerability to nicotine abusing. The availability of transgenic mice expressing the mutant D2 receptor will help establish the causal effect relationship of an up- regulated cAMP pathway and predilection to nicotine abuse. The proposed studies will also lead to an understanding of whether a subtle change in structure of dopamine D2 receptor, one of mediators for the basic neurobiological function of mesolimbic dopamine system, may affect predisposition to nicotine abuse.

[unreadable] DESCRIPTION (provided by applicant): Organization of physiology and behavior with recurring daily environmental conditions is an adaptation that occurs in essentially all living organisms. Circadian rhythms are regulated by three components: the circadian pacemaker, an input mechanism and an output mechanism. In mammals, the master pacemaker driving circadian rhythms resides in the suprachiasmatic nuclei (SCN) of the anterior hypothalamus. Environmental light-dark cycles entrain the SCN clock to the 24-hr day. Synchronization of SCN neurons leads to coordinated circadian outputs that regulate expressed rhythms. A clear view of molecular clock mechanisms within the SCN has emerged. The molecular clockwork of the SCN circadian clock consists of auto-regulatory transcriptional and translational feedback loops that have both positive and negative elements. Similarly, the signal pathway of how light input resets SCN clock to synchronize with the environmental light/dark cycle is also emerging. In contrast to molecular mechanisms of SCN pacemaker clockwork and input pathway, relatively little is known about the output mechanism by which the SCN circadian pacemaker sends timing information to control physiological and behavioral rhythms. Prokineticin 2 (PK2), a cysteine-rich protein, has recently been shown as a SCN output molecule that transmits the circadian locomotor rhythm. PK2 fulfill the criteria expected for a bona fide output molecule from SCN circadian clock: 1) PK2 is a secreted molecule; 2) The transcription of PK2 is regulated by core clock genes, and PK2 mRNA oscillates in the SCN with high magnitude; 3) The production of PK2 is responsive to light entrainment; 4) Receptor for PK2 is expressed in primary SCN output target areas; and 5) Intracerebroventricular (ICV) administration of PK2 at subjective night, when endogenous PK2 levels are low, suppressed high nocturnal wheel-running activity. We propose to further investigate the role of PK2 signaling in the output mechanism of the SCN circadian clock. Specifically, the rhythms of PK2 protein in the cell bodies, terminal areas of SCN neurons and cerebral spinal fluid will be investigated by quantitative immunocytochemistry and/or radioimmunoassay. Whether PK2 is the common signal that mediates the output of SCN circadian clock and light masking will be investigated. How the PK2 rhythmic output from the SCN responds to abrupt shifts of light/dark cycle will also be investigated. Moreover, the effects of PK2 on SCN circadian clock-controlled locomotor and sleep/wake rhythms will be investigated by acute and chronic infusion of PK2 and PK2 antagonist in rats. Furthermore, the PK2 gene will be disrupted in mice and its effect on SCN-controlled circadian behavioral rhythms as well as core SCN circadian loops will be examined. Finally, the SCN PK2 output pathway will be investigated by a genetic approach. These proposed studies should help us gain further insight into the mechanism of PK2 signaling in mediating the output of timing information from the SCN circadian crock, and could have a major impact on the future treatment of a number of circadian disorders such as jet-lag, shift work syndrome, and chronic insomnia [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Despite enormous advances, long term control of hyperglycemia for type 2 diabetes remains a major challenge. Existing pharmacological options of hyperglycemia control include biguanides, sulfonylureas, thiazolidinediones, alpha-glucosidase inhibitors, insulin and incretin agents. These current therapies are often associated with hypoglycemia, weight gain or other adverse events such as gastrointestinal discomfort, edema, cardiac failure or fractures. New therapies that can correct hyperglycemia on a long-term basis without causing adverse events are highly desirable. The success of incretin agents has supported the importance of G-protein coupled receptors on insulin release and therefore glucose homeostasis. Recently, we have accumulated substantial evidence that supports the insulinostatic effect of prokineticin 2 (PK2). PK2, as a secreted regulatory peptide, has previously been shown to regulate diverse biological processes via the activation of two cognate G protein-coupled receptors, prokineticin receptor 1 (PKR1) and prokineticin receptor 2 (PKR2). PK2 as well as PKR1 and PKR2 are expressed in pancreatic beta-cells. PK2-deficient mice have reduced circulating glucose levels, elevated fasting insulin levels, and exhibit enhanced performance in a glucose tolerance assay. Administration of exogenous PK2 was shown to diminish capabilities of glucose clearance in mice. Conversely, administration of a small molecule PK2 receptor antagonist enhances glucose clearance in mice. PK2 decreases glucose-stimulated insulin secretion in acutely isolated pancreatic islets as well as in beta-cell-like MIN6 cells, likely via Gi-coupled pathway. We have also shown that high glucose treatment induces the release of PK2 into conditioned media from MIN6 cells. We have further shown that high fat diet-induced hyperglycemia up-regulates PK2 expression in pancreas islets, indicating dynamic up-regulation of PK2 expression in response to chronic hyperglycemia may disrupt subsequent insulin release in response to glucose load. Taken together, our preliminary findings have indicated that PK2 plays a feedback insulinostatic role on the function of pancreatic beta-cells. We propose to further investigate the role of PK2 signaling on glucose homeostasis. Particularly, we will further examine the glucose-lowering effect of PK2 antagonists in response to oral glucose in lean mice as well as in db/db mice. We will investigate the dynamic change of PK2 expression in pancreas beta-cells in response to high glucose in vitro and in vivo. We will further investigate the role and signaling mechanism of PK2 signaling on insulin release with isolated pancreas islets. Finally, we seek to validate our findings with human subjects that are deficient in PK2 signaling. The successful completion of these studies will lead to the validation of a novel signaling mechanism for insulin release and glucose homeostasis. As one characteristic pathophysiological feature of type 2 diabetes is the inadequate release of insulin from pancreatic beta-cells by glucose, PK2 antagonism via small molecule antagonists to relieve the insulinostatic effect of PK2 may provide a potential novel therapeutic avenue for controlling hyperglycemia. PUBLIC HEALTH RELEVANCE: The prevalence of type 2 diabetes has risen dramatically during recent decades. Due to a variety of adverse effects associated with current pharmacological therapies, control of hyperglycemia for type 2 diabetes remains a major challenge. We propose to examine the role and mechanism of a novel regulatory peptide signaling in insulin release and glucose homeostasis. The successful completion of these studies may lead to the identification of a potential therapeutic strategy and agent for the treatment of type 2 diabetes.