Calcium-dependent autoregulation of vasopressin neurons in a rodent model of heart failure. - PROJECT SUMMARY This K99/R00 proposal will test a critical hypothesis on how vasopressin (VP) neurons self-regulate their neuronal excitability, their local paracrine release of vasopressin, and how these mechanisms are altered in a rodent model of heart failure while providing the PI with additional scientific and professional skills to transition him to a successful independent career as a research professor. Heart failure (HF) is the leading cause of death worldwide. A hallmark of HF is sympathetic hyperactivity and increased circulating levels of neurohormones such as VP. The slow afterhyperpolarization (sAHP) is recognized as a key mechanism that influences VP firing, playing a critical role in shaping the stereotyped phasic bursting patterns in VP neurons. The sAHP is impaired in VP HF neurons. Uniquely, Ca2+-dependent exocytosis of VP occurs somatodendritically (VP-SDR) as well, acting in a paracrine manner as an auto-inhibitory feedback mechanism. It is currently unknown whether VP-SDR is impaired in VP HF neurons. Data presented herein shows sAHPs and VP-SDR share many common signaling pathways which have been observed separately but never in tandem. Furthermore, we have exciting preliminary data that demonstrates VP itself can enhance isolated sAHPs, suggesting that endogenous VP-SDR may modulate sAHP time course, and thus firing activity. Understanding how these mechanisms lead to aberrant VP neuronal activity in HF is of critical importance. I therefore present the unique hypothesis that sAHP and VP-SDR activation are not only activated by the same signaling and pathways, but that VP-SDR can directly modulate sAHPs. I hypothesize that the observed impairment of the sAHP in HF neurons reflects impaired intracellular Ca2+ signaling and VP-SDR, leading to the hyperexcitability and excess peripheral VP release observed in this condition. Aim 1 will test the hypothesis that certain modes of VP activation (ex. continuous vs. phasic stimulation) favors the activation of either sAHPs or VP-SDR. Aim 2 will test the hypothesis that endogenous VP-SDR itself modulates sAHPs. Aim 3 will test the hypothesis that intracellular Ca2+ sensitivity/availability and VP-SDR are impaired in VP neurons of HF rats. This proposal will significantly enhance the PI’s career development and advance him towards his goal of becoming an independent investigator. The proposed project provides training in technical skills and professional development with an emphasis on independence and acquiring skills needed to run a laboratory. We will accomplish this by uniquely integrating previously mastered techniques in a novel way to measure sAHPs and VP-SDR simultaneously via patch clamp and utilization of VP detecting “sniffer” cells, respectively from healthy and HF rats. Georgia State University also provides an exceptional environment for training with the facilities and equipment needed to perform the research, an intellectual environment of seminars and workshops, ongoing collaborations both within and outside the institution, and most critically a mentoring senior PI with a wealth of experience preparing postdocs for an independent career. Indeed, the mentor has provided an explicit plan to actively prepare the PI for independence. In summary, this proposal will provide the training necessary to prepare the applicant for independence while making a significant contribution to our understanding of VP neurons in a prevalent disease.
