The subparaventricular zone (SPZ) of the hypothalamus is an important component of the circuitry that synchronizes and drives circadian rhythms of behavior and physiology. The SPZ receives most of the axonal output from the suprachiasmatic nucleus (SCN), the mammalian circadian pacemaker, and further projects to most of the SCN's downstream targets. Physiological and anatomical studies in rats suggest the SPZ is heterogeneous and comprised of subregions differentially modulating particular rhythms. Specifically, dorsal SPZ lesions, defined by the region extending ventrally from the paraventricular nucleus (PVH), selectively disrupt body temperature (Tb) rhythms with minimal impact on locomotor activity (LMA) or sleep/wake rhythms. In contrast, ventral SPZ lesions, defined by the region extending dorsally and caudally from the SCN, profoundly disrupt LMA and sleep/wake rhythms with less effect on Tb. Anterograde tracing from the SPZ in rats further suggests this area is comprised of four anatomically, and perhaps functionally, distinct subregions: the ventromedial and ventrolateral SPZ (vmSPZ, vlSPZ) and the dorsomedial and dorsolateral SPZ (dmSPZ, dlSPZ). Recent developments of certain conditional knockout mice allow for the direct testing of the neurotransmitter-specific role of SPZ subregions in vivo. We seek to gain a better understanding of SPZ organization and outflow and its role in the circadian timing system in mice. Elucidating such pathways may reveal mechanisms by which chronic circadian disruption alters behavior and adversely affects overall health.
Aim 1. To assess the role of GABAergic subpopulations of the SPZ in the modulation of circadian rhythms of Tb, LMA, and sleep/wake: The SPZ is primarily comprised of GABAergic neurons, which package GABA into vesicles using VGAT. Using Cre-Lox technology, we will inject a viral vector to delete VGAT expression in SPZ subregions, preventing these cells from releasing GABA. We will record freely moving mice in constant dark, while measuring Tb and LMA using biotelemetry transmitters implanted in the intraperitoneal cavity, and sleep/wake using EEG and EMG recordings. We expect these experiments to identify which populations of GABAergic neurons in the SPZ regulate circadian rhythms of LMA, sleep/wake, and Tb.
Aim 2. To delineate efferent projections from GABAergic SPZ neurons and correlate the physiological patterns in Aim 1 with deletion of GABAergic transmission in specific terminal fields: There are no published reports of a detailed analysis of SPZ efferents in mice. The vector used in Aim 1 also traces the efferent projections from transduced neurons and will reveal the projection targets that mediate these rhythms, however, it also traces from non-GABAergic neurons. To clarify the projections likely underlying our effects, we will use mice expressing Cre under the VGAT promoter and a vector expressing the gene for a Cre-dependent tracer to selectively trace SPZ GABAergic projections. We will then compare VGAT deletion in specific targets with physiological responses in Aim 1 to determine SPZ targets regulating specific circadian rhythms.
This proposal focuses on elucidating the neural circuitry involved in synchronizing and driving circadian (daily) rhythms of behavior and physiology. Chronic disruption of these rhythms, such as that which occurs in humans with occupations requiring shift work or frequent travel across time zones, has been associated with numerous negative health outcomes including cardiovascular disease, hypertension, depression, cancer, diabetes and obesity. Thus, a better understanding of the overall circuitry of the circadian timing system may reveal mechanisms for treatment and prevention of some of these deleterious health consequences.