Area-specific reorganization of catecholamine release from C1 and A2 neurones of spontaneously hypertensive rats

Central catecholaminergic (CAergic) neurones, in particular areas C1 and A2, are associated with blood pressure regulation. Links between their activity and central sympathetic outflow have been documented. Since sympathetic drive is elevated in essential hypertension, both in human patients and spontaneously hypertensive rats (SHR), hyperactive central CAergic transmission may be an underlying causative factor. We compared transmitter release characteristics and electrophysiological properties of C1 and A2 neurones of normotensive Wistar rats (WR) with SHR in vitro. Organotypic brainstem slice cultures were transduced with adenoviral vectors to express EGFP specifically in CAergic neurones [1]. CAergic neurones were imaged using a Leica SP confocal microscope and recorded in whole-cell configuration with patch electrodes containing the Ca2+ indicator Rhod-2 [2]. Electrophysiological characteristics of neurones and their intracellular [Ca2+] ([Ca2+]i) responses to angiotensin II (AngII; 200 nM) were determined. For analysis of quantal catecholamine (CA) release characteristics, microamperometry was carried out in separate experiments where carbon fiber microelectrodes were placed onto single varicosities. Oxidation currents were evaluated as in [3]. Microamperometric analysis at C1 release sites in WR and SHR (n=6 each) revealed a striking increase of the relative contribution of large (>0.5 pC) release events in SHR (from 57% to 86% of total CA release) as compared to WR, while contribution of small events (median ~0.025 pC) was diminished (from 43% to 14% of total). This represents a significant redistribution of the mode of release in favour of large quanta in C1 (p<0.05 Chi-square for all fractions). Interestingly, in WR, 48% of C1 neurones (n=44) were spontaneously active while in SHR the majority (90%) were silent (n=10). AngII raised [Ca2+]i in silent C1 neurones of WR and SHR (in 61% and 55%, respectively). This increase was not significantly different between WR and SHR cells (+24 ± 1.8% and +34 ± 8%; p=0.26; Student’s t-test). For area A2, the distribution of various types of release events was similar between WR and SHR. Here, comparable subpopulations of active neurones were found in WR (38%; n=8) and SHR (22%; n=9). For active as well as silent cell groups, the effect of AngII on [Ca2+]i was markedly reduced in SHR (+5 ± 3%) compared to WR (+29 ± 9%; p<0.05, Student’s t-test). In conclusion, in SHR we have uncovered a significant shift in release characteristics of C1 varicosities such that the bulk of CA is released in massive packages in excess of 106 molecules. When released in large packets, transmitter may spread further through extracellular space, opening up a wider range of signalling targets for C1 neurones in SHR. Teschemacher AG et al. (2005) ADDR 57,79-93.