The physiological and cellular actions of hydrogen sulfide on neurons of the subfornical organ

Abstract

Hydrogen sulfide (H2S) is an endogenous gasotransmitter found in the central nervous system (CNS). While difficult to measure, the endogenous concentrations of H2S lie between 10nM and 160μM, and recent work has discovered that H2S exerts beneficial actions in the control of cardiovascular function. In the following thesis, I report data examining the cellular and physiological actions of H2S in the subfornical organ (SFO), a CNS site important in the control of blood pressure. We used male Sprague-Dawley rats for both in vivo and in vitro experiments. In the first manuscript, our RT-PCR experiments confirmed previous microarray analyses showing that mRNAs for H2S-producing enzymes were expressed in the SFO. We then used microinjection techniques to investigate the physiological effects of NaHS in SFO, and found that NaHS microinjection (5nmol) into SFO significantly increased blood pressure (mean area under the curve = 853.5 ± 105.7 mmHg*s, n=5). Furthermore, we used patch clamp electrophysiology and found that 97.8% (88 of 90) of neurons depolarized in response to NaHS in a concentration dependent manner (EC50 of 35.6µM). We also observed an increase in neuronal excitability by analysing the rheobase and action potential firing patterns. In our second manuscript, we used in vitro voltage clamp electrophysiology to study the effects of NaHS on the gating properties of voltage-gated ion channels in the membrane of SFO neurons. We found that NaHS (1mM) depolarized the activation curves of N-type calcium currents, persistent sodium currents, and transient sodium currents by average 50% activation voltage (V50) changes of 5.9mV, 10.8mV, and 7.6mV, respectively. Peak induced currents were also enhanced by NaHS application. Interestingly, we observed no effect on voltage-gated potassium currents. This study has correlated in vivo physiological actions of H2S with in vitro membrane depolarizations and increases in excitability in the SFO, a brain area important to the regulation of cardiovascular function. We have provided evidence of effects of H2S on voltage-gated ion channels contributing to neuronal excitability. This study is the first to identify specific actions of H2S in SFO, and has contributed to the growing literature of H2S actions in the CNS.