Date of Award
Doctor of Philosophy (PhD)
Located in the ventral hypothalamus, the suprachiasmatic nucleus (SCN) is necessary for entraining daily rhythms in physiology and behavior to environmental cues. Though the 20,000 neurons of the SCN uniformly express GABA, they differ greatly in neuropeptide content. One anatomically and functionally distinct class of neuropeptidergic SCN neurons is vasoactive intestinal polypeptide (VIP). Expressed by approximately 10% of SCN neurons, VIP is necessary for synchronizing single-cell SCN rhythms to produce coherent output and sufficient for entrainment. However, little is known about the firing activity of these neurons releases VIP and results in circadian entrainment. We utilized multielectrode array technology and optogenetics to optically tag VIP neurons expressing Channelrhodopsin-2 (ChR2) following three days of spontaneous activity recordings. We find that VIP neurons have circadian firing rates with two distinct patterns, irregular and tonic, that constitute two separate electrophysiological classes. Using optogenetic stimulation in vitro and in vivo, we show that high frequency firing intervals are sufficient to phase shift and entrain circadian rhythms in gene expression and locomotor activity through VIP release. Interestingly, low frequency firing intervals do not phase shift the SCN in vitro and entrain behavioral rhythms more gradually. We also find that stimulation of VIP neurons can only phase delay and entrain rhythms during late subjective day and early subjective night. We conclude that VIP neurons entrain behavior in a time-of-day- and frequency- dependent manner.
Complementary to testing the sufficiency of VIP neuronal firing for entrainment, we tested the necessity of VIP neurons for circadian rhythms in the adult SCN circuit. Using Cre-lox technology in vivo, we triggered adult-onset apoptosis in VIP SCN neurons. We found that over 80% of these mice retained circadian rhythms. We contrast this to Vip null mice, where over 60% lose rhythms. A majority of our mice lacking VIP neurons had decreased locomotor activity periods and increased daily onset variability, which strongly correlated with the intensity of VIP staining. In vitro, deletion of VIP neurons leads to a dramatically reduced amplitude of circadian gene expression and decreases in synchrony on the single-cell level. We conclude that the difference between adult deletion of VIP neurons and Vip null mice suggests a role for VIP in SCN development and in the developed adult circuit VIP neurons are not necessary for rhythmicity.
Finally, we dissected the role of VIP SCN neurons in the daily rhythms in glucocorticoids, by characterizing the anatomy of VIP projections and testing the necessity of VIP neurons. We labeled VIP SCN neurons that project dorsally to the paraventricular nucleus of the hypothalamus (PVN) using a two-color tract tracing experiment. We concluded that a small bilateral subset of VIP SCN neurons projects to each side of the PVN. To test VIP neurons function, we deleted VIP SCN neurons in the adult and measured corticosterone rhythms under constant conditions for 2 days. We find that rhythms in corticosterone are severely dampened with the loss of VIP neurons with peak corticosterone only reaching approximately 50% of wild- type levels. We conclude that VIP SCN neurons contribute stimulatory input to the circadian rhythm in corticosterone.
Taken together, these data suggest that VIP SCN neurons are a heterogeneous class of SCN neurons with multiple roles in adult SCN entrainment, development and the regulation of glucocorticoid rhythms.
Chair and Committee
Erik D. Herzog
Michael Bruchas, Paul Taghert, Tim Holy, John Cirrito,
Mazuski, Cristina, "The Role of VIP SCN Neurons in Circadian Physiology and Behavior" (2017). Arts & Sciences Electronic Theses and Dissertations. 1233.