BRAIN PEPTIDES AND THE REGULATION OF CIRCADIAN RHYTHMS

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The objective of my research program continues to focus on the neural regulation of circadian rhythms by the suprachiasmatic nuclei (SCN) of the hypothalamus. The first set of experiments will analyze the relationship between energy metabolism and function in the SCN using the 2-deoxy-D-(14C)glucose (DG) autoradiographic method. Fetal SCN energy metabolism is rhythmic in utero, before either SCN synaptogenesis or the circadian rhythm of SCN impulse activity becomes apparent post-natally. Experiments will test the hypothesis that the generation & propagation of action potentials by the SCN is not an integral part of the actual circadian timing device. Mini-osmotic pumps will locally perfuse the SCN with one millionth M tetrodotoxin (TTX) at 0.5 Mul/hr for 14 days; free-running rhythmic drinking behavior of blinded rats will be monitored before, during and after infusion of the toxin. This paradigm will be used to (a) uncouple the circadian clock in the SCN from both its afferents and efferents, (b) determine a dose-response relationship for TTX infusion, (c) demonstrate topographical specificity of the agent, and (d) quantitate the resulting rhythm of SCN energy metabolism. The second set of experiments will use the DG method to determine how the maternal circadian system coordinates the timing of the developing fetal clock. Studies for the coming year include (a) maternal SCN ablation at later gestational ages, (b) maternal oophorectomy, and (c) evaluation of non-endocrine behavioral mechanisms using food restriction, shifting of mealtimes, and continuous intravenous hyperalimentation regimes. The third set of experiments will investigate the physiological role for the circadian vasopressin rhythm in cerebrospinal fluid (CSF). Preliminary data suggest that the circadian rhythm of brain temperature is severely disturbed in homozygous Brattleboro rats. Further characterization of this abnormality will lead to experiments to (a) abolish the normal temperature rhythms of Long-Evans rats by intracerebroventricular administration of vasopressin antibody or antagonist, and (b) restore temperature rhythmicity in Brattleboro homozygotes by artificial simulation of normal CSF vasopressin rhythms or by transplantation of the fetal SCN. Vasopressin is measured by a sensitive and specific radioimmunoassay.