Jaya Pamidimukkala

Expression of metabotropic glutamate receptor 8 in autonomic cell groups of the medulla oblongata of the rat.

Metabotropic glutamate receptors (mGluRs) in the medulla oblongata have been suggested to have a functional role in the regulation of cardiovascular baroreflexes. The present study examines the localization of mGluR8 autonomic nuclei of the medulla of the rat. mGluR8 immunoreactivity was observed in the cell bodies and/or processes of the dorsolateral, interstitial, medial, intermediate, ventral, ventrolateral, subpostremal, commissural, parvicellular and gelatinosus subnuclei of the nucleus tractus solitarius (NTS). The intensity of mGluR8 staining was highest in the commissural and interstitial subnuclei at the level of the area postrema. Commissural NTS is involved in regulation of baro-, and chemo-reflexes whereas the interstitial nucleus mediates respiratory reflexes. In the area postrema, diffuse staining was observed in the cell bodies, dendrites and fibers of the dorsal and central regions. In vagal outflow nuclei, mGluR8 immunoreactivity was observed in: (1). the cell bodies and processes of the dorsal motor nucleus of the vagus (DMN) throughout the rostro-caudal extent; and (2). the cell bodies and fibers throughout the rostro-caudal extent of the dorsal and ventral division the nucleus ambiguus (NA). Staining in the ventrolateral medulla was restricted to regions ventral to the nucleus ambiguus and dorsal to the lateral reticulate nucleus. The present study is the first to provide a detailed mapping of mGluR8 within the autonomic nuclei of the medulla and suggests that this subtype may be involved in shaping synaptic transmission in these central nuclei.

Cellular Mechanisms Regulating Synaptic Vesicle Exocytosis and Endocytosis in Aortic Baroreceptor Neurons

Abstract: The purpose of this chapter is to review some of the recent progress in the understanding of the cellular and biophysical mechanisms that are involved in the regulation of arterial baroreceptor neurotransmssion. Synaptic depression or fatigue following repeated neuronal stimulation has been shown at central baroreceptor synapses in vivo and in vitro. As most of the central neurons have a limited number of vesicles, vesicle retrieval or endocytosis following exocytosis is thought to play a major role in preserving synaptic transmission. We have hypothesized that central baroreceptor terminals may inhibit their own synaptic transmission via feedback activation of presynaptic metabotropic glutamate receptors (mGluRs). We have analyzed the effects of mGluR autoreceptors (group III mGluRs) on voltage‐gated calcium channels using standard patch‐clamp techniques and on the process of exocytosis and endocytosis in aortic baroreceptor neurons using the quantitative imaging dye FM1–43 and FM2–10. Usng the whole‐cell patch‐clamp technique, we have found that activation of group III mGluRs with L‐AP4 inhibits peak calcium channel current. Furthermore, activation of group III mGluRs with L‐AP4 markedly decreases stimulation‐induced exocytosis in aortic baroreceptor neurons, as measured with FM1–43, and inhibits synapsin I phosphorylation. These results suggest that activation of group III mGluRs may inhibit synaptic transmission by (1) inhibiting calcium influx, (2) decreasing synaptic vesicle exocytosis, and (3) modulating the mechanisms governing synaptic vesicle recovery and endocytosis. These effects of mGluRs on baroreceptor synaptic vesicles may contribute to the baroreceptor/nucleus tractus solitarius synaptic depression observed in vivo.

Effects of salt rich diet in the obese Zucker rats: studies on renal function during isotonic volume expansion.

Obese Zucker rats (OZR) are hyperinsulenemic, hyperglycemic and dyslipidemic and develop salt dependent hypertension. Since salt sensitivity is considered to be due to impaired handling of renal sodium excretion, these studies were conducted in the obese and lean Zucker rats (LZR) anesthetized with Inactin to evaluate renal function under basal conditions and during acute isotonic fluid volume expansion (VE). Mean Arterial blood pressure (MBP), heart rate (HR), renal blood flow(RBF) and glomerular filtration rate (GFR) were not significantly different between the lean Zucker rats fed normal diet or that fed salt rich diet(8% NaCl). However, basal UV and UNaV were significantly greater in the LZR fed high salt. During VE essentially identical increases occurred in GFR, UV and UNaV in both the lean groups. In the OZR fed salt rich diet also, there were no significant changes in the heart rate, RBF and GFR. However, arterial blood pressure of the OZR fed salt rich diet was significantly greater than that of the OZR on the normal diet as well as that of both the lean groups. Also, as in the LZR, basal UV and UNaV were significantly greater in the salt fed obese rats. During volume expansion there were no impairments in the ability of the obese groups fed normal or salt rich diet to eliminate sodium and water during volume load. In fact, the net sodium and water excretions during and 60 min after VE in both the obese groups were significantly greater than that of corresponding lean groups. Furthermore, these values in the OZR fed salt rich diet were significantly greater than that of the obese rats on normal salt diet perhaps due to the contribution of ‘pressure natriuretic mechanisms’. These data demonstrate that although OZR are salt sensitive, the renal mechanisms that would collectively respond to acute isotonic VE were fully functional. An unexpected and a novel finding in these studies is that the salt rich diet, in addition to increasing arterial blood pressure also significantly lowered plasma of insulin levels and enhanced glucose and cholesterol levels in the obese Zucker rats.

Frequency dependence of synaptic vesicle exocytosis in aortic baroreceptor neurons and the role of group III mGluRs.

Synaptic transmission between baroreceptor afferents and the nucleus tractus solitarius (NTS) is essential for reflex regulation of blood pressure. High frequency stimulation of the afferents in vivo leads to a decrease in synaptic strength and is generally attributed to reduction in presynaptic neurotransmitter release. It has been hypothesized that during high frequency stimulation glutamate a major neurotransmitter at the baroreceptor afferent terminals inhibits its own release via presynaptic group III metabotropic glutamate receptors (mGluRs). A key player in modulation of presynaptic release is vesicle exocytosis. The present study utilized cultured aortic baroreceptor neurons and the styryl dye FM2-10 to characterize (1) the dependence of exocytosis at these afferent nerve terminals on the frequency of neuronal activation, (2) the effect of duration of stimulation on the rate of exocytosis and (3) the role of mGluRs in the frequency-dependent modulation of exocytosis. Destaining in the FM2-10 loaded boutons during 3 min of stimulation, a measure of exocytosis, progressively decreased with increasing frequency (0.5, 1.0 and 10 Hz). Blockade of group III mGluRs with 300 microM (RS)-cyclopropyl-4-phosphonophenylglycine (CPPG) facilitated exocytosis evoked by 10 Hz stimulation but not at 0.5 Hz. The data suggest that aortic baroreceptor terminals exhibit frequency-dependent depression of exocytosis and support a role for group III mGluRs in the frequency-dependent modulation of exocytosis.

Frequency-dependent depression of exocytosis and the role of voltage-gated calcium channels

Synaptic vesicle exocytosis in primary cultures of baroreceptor neurons is reduced during high-frequency stimulation. Calcium influx through voltage-gated calcium channels (VGCC) is a key step in neurotransmitter release. With the help of FM2-10, a marker of synaptic vesicle recycling, the present study investigates the differential contribution of several VGCC subtypes to exocytosis in neuronal processes and how this contribution is altered at high frequencies. In control experiments, field stimulation at 0.5 Hz evoked about 66 +/- 5% destaining. Combined blockade of N- and P/Q-subtypes with Ctx-MVIIC was far more effective in reducing exocytosis (11 +/- 8%) than blocking N-type (49 +/- 5%, Ctx-GVIA) or P-type (46 +/- 1%, Agatoxin) alone. The effectiveness of the blockers also varied with the duration of stimulation: Ctx-GVIA attenuating exocytosis significantly in the first 60 s and Agatoxin affecting exocytosis only towards the end of 180 s stimulation period. Field stimulation at 10 Hz evoked exocytosis (36 +/- 18%) comparable to that evoked by 0.5 Hz in the presence of Ctx-GVIA. While blockade with Agatoxin had no effects, Ctx-GVIA, Ctx-MVIIC and L-type blocker Nifedepine had small but similar inhibitory effects on exocytosis at 10 Hz. The data suggest that N-type is the major contributor to exocytosis at 0.5 Hz, and this contribution is reduced during prolonged stimulation periods and at high frequencies.