Salt-induced hypertension: role of purinergic signaling in the neuronal cells at the hypothalamus level, and its correlation with the autonomic nervous system and blood pressure control

Abstract

The main purpose of this project is to evaluate the mechanisms of purinergic neurotransmission between neuronal and glial cells of the paraventricular hypothalamic nucleus (PVN), whose changes in their neurochemistry directly involved in the control the autonomic nervous system (ANS), and the genesis of salt-induced hypertension. High salt intake leads to increase in the plasma osmolality, and consequently raise sympathetic nerve activity (SNA) by mechanisms that involve hypothalamic neuronal activation, particularly PVN, which contributes to neurogenic hypertension induced by high salt intake. Recently, our laboratory have shown that adenosine triphosphate (ATP) increases the activity of neurons in the PVN, and causes an increase of SNA via activation of purinergic receptors, P2 subtype, and a co-transmission with glutamatergic pathways. Thus, our hypothesis in this project is that hypertension and sympathoexcitation elicited by high salt intake depends on the purinergic signaling at the PVN level, and it involves a neuron/glia and ectonucleotidase interactions responsible for the availability of this purine at neuronal microenvironment of this nucleus. In turn, this phenomenon could be due a higher release of this purine by glial cells, which is consider to be a putative source of it, or at least a reduction of ectonucleotidases' activity. Male Wistar rats will be submitted to salt overload (NaCl 2%) to evaluate the expression of FOS protein and its co-location with P2X2 receptors in the PVN. In addition, it will be evaluated whether the P2 receptors participate in sympathoexcitatory responses to salt loading. Additionally, we purpose to evaluate if hyperosmotic stimulus is able to promote release of ATP within the PVN, by using biosensor that detects the release of this purine in real time. To evaluate the role of glial cells in ATP release, PVN cell cultures will be stimulated with hypertonic solution in the presence of fluoroacetate, a glial activity inhibitor. PCR technique will be performed to assess whether the salt loading modify the mRNA expression of enzymes ectonucleotidase (E-NTPDase1,2,3,8) in the PVN. The function of ectonucleotidades will be evaluated in vivo by using gene therapy techniques with lentivirus vector (LVV) encode transmembrane prostate acidic phosphatase (TMPAP) called of LVV-EF1a-TMPAP-EGFP to increase the expression of these enzymes at the PVN neuronal level. In this context, changes in cardiovascular parameters, blood pressure and heart rate will be evaluated in non-anesthetized freely moving animals via radiotelemetry technique. (AU)