Meredith Hay

Mineralocorticoid excess increases superoxide production by activating NADPH oxidase (NOX), and intracerebroventricular infusions of NADPH oxidase inhibitors attenuate aldosterone (Aldo)/salt-induced hypertension. It has been hypothesized that increased reactive oxygen species (ROS) in the brain may be a key mechanism in the development of hypertension. The present study investigated the brain regional specificity of NADPH oxidase and the role of NOX2 and NOX4 NADPH oxidase subunits in the hypothalamic paraventricular nucleus (PVN) in Aldo/salt-induced hypertension. PVN injections of adenoviral vectors expressing small interfering (si)RNA targeting NOX2 (AdsiRNA-NOX2) or NOX4 (AdsiRNA-NOX4) mRNAs were used to knock down NOX2 and NOX4 proteins. Three days later, delivery of Aldo (0.2 mg·kg(-1)·day(-1) sc) via osmotic pump commenced and 1% NaCl was provided in place of water. PVN injections of either AdsiRNA-NOX2 or AdsiRNA-NOX4 significantly attenuated the development of Aldo/NaCl-induced hypertension. In an additional study, Aldo/salt-induced hypertension was also significantly attenuated in NOX2 (genomic) knockout mice compared with wild-type controls. When animals from both functional studies underwent ganglionic blockade, there was a reduced fall in blood pressure in the NOX2 and NOX4 knockdown/knockout mice. Western blot analyses of the PVN of siRNA-NOX2- or siRNA-NOX4-injected mice confirmed a marked reduction in the expression of NOX2 or NOX4 protein. In cultured PVN neurons, silencing either NOX2 or NOX4 protein production by culturing PVN cells with siRNA-NOX2 or siRNA-NOX4 attenuated Aldo-induced ROS. These data indicate that both NOX2 and NOX4 in the PVN contribute to elevated sympathetic activity and the hypertensivogenic actions induced by mineralocorticoid excess.

Serotonergic fibers have been identified within the granule cell layer of the cerebellar cortex; however, their functional significance has not been identified. In this study the effect of serotonin on granule cell spontaneous activity was determined in the rat cerebellum. Of the 136 granule cells tested, 44.8% displayed a decrease in firing rate, 21.3% increased firing rate and 33.8% were not affected. The serotonin-induced changes in activity were not blocked by bicuculline or methysergide. The serotonin agonist 1,3 (trifluoromethylphenyl) piperazine mimicked the serotonin-induced suppressive response. Iontophoretically applied serotonin was also found to modulate GABA-induced suppression of granule cell activity. The variable effects of serotonin on spontaneous activity suggests the presence of more than one type of serotonergic receptor in the cerebellar granule cell layer.

The nucleus tractus solitarius (NTS) receives information from both area postrema (AP) and peripheral afferents. It is, therefore, one likely site of interaction between AP and peripheral afferent fibers. The present study's purpose was to determine the influence of AP stimulation on solitary tract-induced modulation of NTS neuronal activity. With the use of an in vitro rabbit brain slice preparation, extracellular recordings were made from 58 NTS neurons in which action potentials were evoked by both solitary tract and AP stimulation. In the majority of the cells tested, simultaneous stimulation of solitary tract and AP, at voltage levels that evoked no action potentials when stimulated separately, resulted in production of either single or multiple action potentials. In 27 units, stimulation levels to the solitary tract and to the AP were adjusted such that their respective separate stimulations produced an NTS action potential less than 30% of the time. When the two inputs were stimulated together, simultaneous stimulations produced an NTS action potential 100% of the time, suggesting a facilitatory interaction between the AP and the solitary tract on NTS neuronal activity. In nine cells, perfusion of the slice with clonidine induced a facilitation of solitary tract-evoked NTS response to a level similar to that seen during simultaneous stimulation of the solitary tract with the AP. Application of the alpha 2-adrenergic receptor antagonist yohimbine blocked the ability of both clonidine and AP to facilitate the solitary tract-evoked response. These results support a possible interaction between AP and peripheral afferents and suggest that AP stimulation facilitates effects of solitary tract activation at the level of the NTS.

Glutamate is the proposed neurotransmitter of baroreceptor afferents at the level of the nucleus of the solitary tract (NTS). Blockade of ionotropic glutamate receptors with kynurenic acid blocks the arterial baroreflex but, paradoxically, does not abolish the response to exogenous glutamate. This study tested the hypothesis that exogenous glutamate in the NTS activates both ionotropic and metabotropic glutamate receptors (mGluRs). In urethan-anesthetized rats, unilateral microinjections of glutamate into the NTS decreased mean arterial pressure, heart rate, and lumbar sympathetic nerve activity. The cardiovascular response to injection of glutamate was not altered by NTS blockade of mGluRs with alpha-methyl-4-carboxyphenylglycine (MCPG). Blockade of ionotropic glutamate receptors with kynurenic acid attenuated the response to glutamate injection. After combined NTS injection of MCPG and kynurenic acid, the response to glutamate was blocked. These data suggest that exogenous glutamate microinjected into the NTS acts at both ionotropic glutamate receptors and mGluRs. In addition, blockade of both classes of glutamate receptors is required to block the cardiovascular response to microinjection of glutamate in the NTS.

1. Calcium currents in rabbit area postrema neurons were studied with the perforated patch-clamp technique. Experimental conditions eliminated Na+ and K+ currents and identified both low- and high-threshold voltage-activated calcium currents. 2. Low-threshold, T-type calcium currents were observed in 64% of the area postrema neurons recorded. This current activated near -60 mV and had an average peak amplitude of -36.2 +/- 5 pA (mean +/- SE) at -40 mV. This current began rapid inactivation near -95 mV, reached half-maximal inactivation at -71 mV and was totally inactivated by -40 mV. 3. A high-threshold transient current was recorded in all area postrema neurons, which consisted of both a transient and sustained component. This current was present at voltages greater than -40 mV and the transient component of this current was responsible for the majority of the total Ca2+ current. 4. Nickel ions (10 microM) effectively reduced both the T-type current and the high-threshold current. Cadmium ions (100 microM) effectively reduced the high-threshold current while having insignificant effects on the low-threshold current. 5. Application of the dihydropyridine antagonist nimodipine (1-10 microM) had no effect on either the low- or high-threshold voltage-activated calcium Ca2+ in area postrema neurons. In addition, application of omega-conotoxin-GVIA (2-10 microM) was also without effect on either the low- or high-threshold voltage-activated Ca2+ current, suggesting that area postrema neurons possess neither L- or N-type voltage-activated Ca2+ currents. 6. Application of omega-conotoxin MVIIC (10 microM) significantly inhibited the peak high-threshold Ca2+ current by 65.4% suggesting that area postrema neurons do possess a omega-conotoxin MVIIC-sensitive high-threshold Ca2+ channel. 7. Arg-vasopressin (150 nM) significantly increased the transient component of the high-threshold Ca2+ current but had little effect on either the low-threshold or the high-threshold sustained component.