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Adrenaline (Adr) is known to directly or indirectly modulate bone cell activity under physiological and pathological conditions. Osteoblasts play a major role in bone formation, employing intracellular Ca(2+) as a second messenger to modulate hormonal responses and as a cofactor for mineralization. Voltage-dependent Ca(2+) channels (VDCCs) mediate the influx of Ca(2+) in response to membrane depolarization. The purpose of this study was to investigate the effects of Adr on VDCC currents in osteoblasts using a patch-clamp recording method. Application of 1 mM Adr facilitated VDCC currents in a concentration-dependent manner. Pre-treatment with b receptor antagonist propranolol blocked Adr-induced facilitation of VDCC currents carried by Ba(2+) (IBa). These results indicate that Adr-induced facilitation of IBa was mediated by b receptors in MC3T3-E1 osteoblast-like cells. To our knowledge, the data presented here demonstrate for the first time that Adr facilitates VDCCs in MC3T3-E1 osteoblast-like cells.

The hypothalamic nonapeptide and neurohypophyseal hormone arg-vasopressin (AVP), also known as antidiuretic hormone, is best known for its effects on water reabsorption in kidney. Osteoblasts play a major role in bone formation, employing intracellular Ca(2+) as a second messenger to modulate hormonal responses and as a cofactor for mineralization. Voltage-dependent Ca(2+) channels (VDCCs) mediate the influx of Ca(2+) in response to membrane depolarization. The purpose of this study was to investigate the effects of AVP on VDCC currents in osteoblasts using a patch-clamp recording method. An application of 1μM AVP facilitated VDCC currents in osteoblasts. To our knowledge, the data presented here demonstrate for the first time that AVP facilitates VDCCs in osteoblasts.

Angiotensin II (Ang II) plays a major role in the maintenance of extracellular fluid volume and blood pressure. In addition to its well-established role in circulatory homeostasis, it has been implicated in the process of bone formation. Osteoblasts play a major role in bone formation, employing intracellular Ca(2+) as a second messenger to modulate hormonal responses and as a cofactor for mineralization. Voltage-dependent Ca(2+) channels (VDCCs) mediate the influx of Ca(2+) in response to membrane depolarization. The purpose of this study was to investigate the effects of Ang II on VDCC currents in osteoblasts using a patch-clamp recording method. To our knowledge, the data presented here demonstrate for the first time that Ang II facilitates VDCCs in osteoblasts.

It is established that neuropeptide Y (NPY) is a transmitter of parasympathetic secretory impulses in submandibular gland. The neuropeptides substance P, vasoactive intestinal peptide (VIP) and calcitonin gene-related peptide (CGRP) are likely mediators of secretory parasympathetic responses of the gland. Previously, we have shown that substance P, VIP and CGRP modulate voltage-dependent Ca(2+) channels (VDCCs) in hamster submandibular ganglion (SMG) neurons. In this study, we attempt to characterize the effect of NPY on VDCCs current using Ba(2+) (I(Ba)) in SMG neurons. Application of NPY caused both facilitation and inhibition of L-type and N/P/Q-type I(Ba), respectively. Intracellular dialysis of the Gα(s)-protein antibody attenuated the NPY-induced facilitation of I(Ba). The adenylate cyclase (AC) inhibitor, as well as protein kinase A (PKA) inhibitor attenuated the NPY-induced facilitation of I(Ba). Intracellular dialysis of the Gα(i)-protein antibody attenuated the NPY-induced inhibition of I(Ba). Application of a strong depolarizing voltage prepulse attenuated the NPY-induced inhibition of I(Ba). These results indicate that NPY facilitates L-type VDCCs via Gα(s)-protein involving AC and PKA. On the other hand, NPY also inhibits N/P/Q-type VDCCs via Gα(i)-protein βγ subunits in the SMG neurons., (Copyright © 2012 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.)

Osteoblasts play a major role in bone formation. Osteoblasts employ intracellular Ca(2+) as a second messenger modulating hormonal responses and a cofactor for bone mineralization. Voltage-dependent Ca(2+) channels (VDCCs) are most commonly present in excitable cell membranes. They are also present at lower levels even in most nonexcitable cells too. In both types of cell, they mediate the influx of Ca(2+) in response to membrane depolarization. Prepulse facilitation is a phenomenon in which a long and strong depolarizing pulse induces a form of VDCC that exhibits an increased opening probability. We believe this to be the first study to demonstrate that strong depolarization prepulses both increase and decrease VDCCs in osteoblasts.

Objective: The control of saliva secretion is mainly under parasympathetic control. The submandibular ganglion (SMG) is a parasympathetic ganglion which receives inputs from preganglionic cholinergic neurons, and innervates the submandibular salivary gland to control saliva secretion. The aim of this study was to investigate if adrenomedullin (ADM) and/or calcitonin gene-related peptide (CGRP) modulate voltage-dependent calcium channel (VDCCs) current (I(Ca)) in SMG., Design: The profile of CGRP and ADM actions in SMG was studied using the whole-cell configuration of the patch-clamp technique., Results: Both ADM and CGRP facilitated I(Ca). These facilitations were attenuated by intracellular dialysis of the anti-Gα(s)-protein and pretreatment of SQ22536 (an adenylate cyclase inhibitor)., Conclusions: ADM and CGRP facilitates VDCCs mediated by Gα(s)-protein and adenylate cyclase in SMG., (Copyright © 2010. Published by Elsevier Ltd.)

The biologically active form of vitamin D, 1α,25-dihydroxy vitamin D₃ (VD), regulates the synthesis of the bone Ca-binding proteins osteocalcin and osteopontin. The actions of VD are mediated through the vitamin D receptor (VDR). Liganded VDR heterodimerizes with the retinoid X receptor and interacts with a vitamin D response element (VDRE). Recently, it has been demonstrated that vitamin D responses elicited in osteoblasts can be rapid as well as long-term. The purpose of this study was to elucidate the mechanism of Ca(2+) signaling of VD in osteoblasts using intracellular Ca(2+) ([Ca(2+)]i) measurements. A rapid VD (10 nM)-induced increase in [Ca(2+)]i was observed within 40 sec. This increase, however, was negated with application of Ca(2+)-free Krebs' solution. These results indicate that VD induces an increase in [Ca(2+)]i from extracellular Ca(2+) in osteoblasts.

Objective: Neurotrophins, such as nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF), promote neuronal development and neuronal survival, but their mechanisms remain controversial. This study aimed to investigate the hypothesis that NGF and BDNF interfere with angiotensin-II- and glutamate-induced facilitation of voltage-dependent Ca(2+) channels (VDCCs) in nucleus tractus solitarius (NTS) neurons., Design: The profile of NGF and BDNF actions in acutely dissociated rat NTS was studied using the whole-cell configuration of the patch-clamp technique., Results: Pretreatment with NGF and BDNF attenuated angiotensin-II-induced facilitation of VDCCs, but did not attenuate glutamate-induced facilitation of the L-type VDCC current in NTS neurons. NGF-induced attenuation was antagonised by pretreatment with a tyrosine kinase A (TrkA) receptor antagonist K-252a., Conclusions: NGF attenuated angiotensin-II-induced facilitation of L-type VDCCs mediated by TrkA receptors in NTS neurons.

Galanin (GAL), a 29-amino-acid neuropeptide, is involved in various neuronal functions, including the regulation of food intake, hormone secretion and central cardiovascular regulation. The nucleus tractus solitarius (NTS) is known to plays a major role in the regulation of cardiovascular, respiratory, gustatory, hepatic and swallowing functions. Voltage-dependent Ca2+ channels (VDCCs) serve as crucial mediators of membrane excitability and Ca(2+)-dependent functions such as neurotransmitter release, enzyme activity and gene expression. The purpose of this study was to investigate the effects of GAL on VDCCs currents (ICa) carried by Ba2+ (IBa) in the NTS using patch-clamp recording methods. An application of M617 (GalR1 specific agonist), AR-M961 (GAL receptor GalR 1/2 agonist) and GAL caused inhibition of N- and P/Q-types I(Ba). M617, GAL, and AR-M961 caused inhibition of I(Ba) in a concentration-dependent manner, with IC50s of 678 nM, 325 nM and 573 nM, respectively. This inhibition was relieved, albeit incompletely, by a depolarizing prepulse. Pretreatment with M35 (GalR non-specific antagonist) attenuated the M617-induced inhibition of I(Ba). Intracellular dialysis of the Galpha(i)-protein antibody also attenuated the Gal-induced inhibition of IBa. These results indicate that GAL inhibits N- and P/Q-types VDCCs via Galpha(i)-protein betagamma subunits mediated by GalR1 in NTS.

Neurokinins, such as substance P (SP), modulate the reflex regulation of cardiovascular and respiratory function in the CNS, particularly in the nucleus tractus solitarius (NTS). There is considerable evidence of the action of SP in the NTS, but the precise effects have not yet been determined. Voltage-dependent Ca2+ channels (VDCCs) serve as crucial mediators of membrane excitability and Ca2+ -dependent functions such as neurotransmitter release, enzyme activity and gene expression. The purpose of this study was to investigate the effects of neurokinins on VDCCs currents (ICa) in the NTS using patch-clamp recording methods. In 142 of 282 neurons, an application of [Sar(9), Met(O(2)11]-substance P (SSP, NK(1) receptor agonist) caused facilitation of L-type I(Ba). Intracellular dialysis of the Galpha(q/11)-protein antibody attenuated the SSP-induced facilitation of I(Ba). In addition, phospholipase C (PLC) inhibitor, protein kinase C (PKC) inhibitor and PKC activator attenuated the SSP-induced the facilitation of I(Ba). In contrast, in 115 of 282 neurons, an application of SSP caused inhibition of N- and P/Q-types I(Ba). Intracellular dialysis of the Gbetagamma-protein antibody attenuated the SSP-induced inhibition of I(Ba). These results indicate that NK(1) receptor facilitates L-type VDCCs via Galpha(q/11)-protein involving PKC in NTS. On the other hand, NK(1) receptor inhibits N- and P/Q-types VDCCs via Galpha(q/11)-protein betagamma subunits in NTS.

1. The profile of opioid and cannabinoid receptors in neurons of the nucleus tractus solitarius (NTS) has been studied using the whole-cell configuration of the patch clamp technique. 2. Experiments with selective agonists and antagonists of opioid, ORL and cannabinoid receptors indicated that mu-opioid, kappa-opioid, ORL-1 and CB1, but not delta-opioid, receptors inhibit VDCCs in NTS. 3. Application of [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO; mu-opioid receptor agonist), Orphanin FQ (ORL-1 receptor agonist) and WIN55,122 (CB1 receptor agonist) caused inhibition of I(Ba) in a concentration-dependent manner, with IC50's of 390 nM, 220 nM and 2.2 microM, respectively. 4. Intracellular dialysis of the G(i)-protein antibody attenuated DAMGO-, Orphanin FQ- and WIN55,122-induced inhibition of I(Ba). 5. Both pretreatment with adenylate cyclase inhibitor and intracellular dialysis of the protein kinase A (PKA) inhibitor attenuated WIN55,122-induced inhibition of I(Ba) but not DAMGO- and Orphanin FQ-induced inhibition. 6. Mainly N- and P/Q-type VDCCs were inhibited by both DAMGO and Orphanin FQ, while L-type VDCCs were inhibited by WIN55,122. 7. These results suggest that mu- and kappa-opioid receptors and ORL-1 receptor inhibit N- and P/Q-type VDCCs via G alpha(i)-protein betagamma subunits, whereas CB1 receptors inhibit L-type VDCCs via G alpha(i)-proteins involving PKA in NTS.

It is recognized that brain contains all the components of the renin-angiotensin systems (RAS). The nucleus of the tractus solitarius (NTS) is known to play a major role in the regulation of cardiovascular, respiratory, gustatory, hepatic and swallowing functions. Voltage-dependent Ca2+ channels (VDCCs) serve as crucial mediators of membrane excitability and Ca2+-dependent functions such as neurotransmitter release, enzyme activity and gene expression. The purpose of this study was to investigate the effects of angiotensin II (Ang II) on VDCC currents (I(Ca)) in the NTS using patch-clamp recording methods. An application of Ang II caused facilitation of L-type I(Ca) in a concentration-dependent manner with an EC50 of 167 nm and a Hill coefficient of 1.73. AT1 receptor antagonist losartan antagonized the Ang II-induced facilitation of I(Ca). Intracellular dialysis of the Galpha(i)-protein antibody attenuated the Ang II-induced facilitation of I(Ca). Both Src tyrosine kinase inhibitor and mitogen-activated protein kinase (MAPK) inhibitor attenuated the Ang II-induced facilitation of I(Ca). p38 MAPK inhibitor also attenuated the Ang II-induced facilitation of I(Ca). These results indicate that Ang II facilitates L-type VDCCs via Galpha(i)-proteins involving Src tyrosine kinase and p38 MAPK kinase mediated by AT1 receptors in NTS.

It is well known that metabotropic glutamate receptors (mGluRs) have multiple actions on neuronal excitability mediated by G-protein-coupled receptors, although the exact mechanisms by which these actions occur are not understood. This study examines the effects of mGluRs agonists on voltage-dependent Ca2+ channels (VDCCs) currents (ICa) in the nucleus tractus solitarius (NTS) of rats using patch-clamp recording methods. An application of (RS)-3,5-dihydroxyphenylglycine (DHPG, Group I mGluR agonist) caused both facilitation and inhibition of L-type and N/P/Q-types ICa, respectively. Neither (2S, 2'R, 3'R)-2-(2', 3'-dicarboxycyclopropyl)glycine (DCG, Group II mGluRs agonist) nor L-(+)-2-amino-4-phosphonobutyric acid (AP-4, Group III mGluRs agonist) nor (RS)-2-chloro-5-hydroxyphenylglycine (CHPG, mGluR5 agonist) modulated ICa. Intracellular dialysis of the Gq/11-protein antibody and Gi-protein antibody attenuated the DHPG-induced facilitation and inhibition, respectively. The phospholipase C (PLC) inhibitor, as well as inhibition of either the protein kinase C (PKC) or inositol-1,4,5-trisphosphate (IP3) attenuated the DHPG-induced facilitation of ICa but not a DHPG-induced inhibition. Application of a strong depolarizing voltage prepulse attenuated the DHPG-induced inhibition of ICa. These results indicate that mGluR1 facilitates L-type VDCCs via Gq/11-protein involving PKC including IP3 formation. On the other hand, mGluR1 inhibits N- and P/Q-types VDCCs via Gi-protein betagamma subunits.

The control of saliva secretion is mainly under parasympathetic control, although there also could be a sympathetic component. Sympathetic nerves are held to have a limited action in secretion in submandibular glands because, on electrical stimulation, only a very small increase to the normal background, basal secretion occurs. Parasympathetic stimulation, on the other hand, caused a good flow of saliva with moderate secretion of acinar mucin, plus an extensive secretion of granules from the granular tubules. The submandibular ganglion (SMG) is a parasympathetic ganglion which receives inputs from preganglionic cholinergic neurons, and innervates the submandibular salivary gland to control saliva secretion. Neurotransmitters and neuropeptides acting via G-protein coupled receptors (GPCRs) change the electrical excitability of neurons. In these neurons, many neurotransmitters and neuropeptides modulate voltage-dependent calcium channels (VDCCs). The modulation is mediated by a family of GPCRs acting either directly through the membrane delimited G-proteins or through second messengers. However, the mechanism of modulation and the signal transduction pathway linked to an individual GPCRs depend on the animal species. This review reports how neurotransmitters and neuropeptides modulate VDCCs and how these modulatory actions are integrated in SMG systems. The action of neurotransmitters and neuropeptides on VDCCs may provide a mechanism for regulating SMG excitability and also provide a cellular mechanism of a variety of neuronal Ca(2+)-dependent processes.

The Vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) are two novel neuropeptides which produce particular biological effects caused by interaction with G-protein-coupled receptors. We have shown in a previous study where VIP and PACAP 38 inhibit voltage-dependent calcium channel (VDCC) currents (ICa) via G-proteins in hamster submandibular ganglion (SMG) neurons. In this study, we attempt to further characterize the signal transduction pathways of VIP-and PACAP 38-induced modulation of ICa. Application of 1 microM VIP and PACAP 38 inhibited ICa by 33.0 +/- 3.1% and 36.8 +/- 2.6%, respectively (mean +/- S.E.M., n = 8). Application of strong voltage prepulse attenuated PACAP 38-induced inhibition of ICa. Pretreatment of cAMP dependent protein kinase (PKA) activator attenuated VIP-induced inhibition, but not the PACAP 38-induced inhibition. Intracellular dialysis of the PKA inhibitor attenuated the VIP-induced inhibition, but not the PACAP 38-induced inhibition. Pretreatment of protein kinase C (PKC) activator and inhibitor attenuated VIP-induced inhibition, but not the PACAP 38-induced inhibition. Pretreatment of cholera toxin (CTX) attenuated PACAP 38-induced inhibition of ICa. These findings indicate that there are multiple signaling pathways in VIP and PACAP 38-induced inhibitions of ICa: one pathway would be the VPAC1/VPAC2 receptors-induced inhibition involving both the PKA and PKC, and another one concerns the PAC1 receptor-induced inhibition via Gs-protein betagamma subunits. The VIP-and PACAP 38-induced facilitation of ICa can be observed in the SMG neurons in addition to inhibiting of ICa., (Copyright 2004 Elsevier B.V.)

The postsynaptic actions of exogenously applied adenosine 5'-triphosphate (ATP) were investigated in nucleus tractus solitarius (NTS) of the rat. Whole cell patch-clamp recordings were used to examine the regulation of voltage-dependent Ca2+ channels (VDCCs) currents (I(Ca)) by ATP in freshly dissociated NTS. Application of ATP inhibited I(Ca) from -905 pA to -741 pA. In addition to this inhibition, application of ATP facilitated I(Ca) from -941 pA to -1,094 pA in other neurons. The data presented here demonstrate for the first time that ATP has both inhibitory and facilitative effects on I(Ca) in NTS. It can be considered that ATP acts as a neurotransmitter in the NTS by having multiple regulatory effects on VDCCs.

1. The presence and profile of purinoceptors in neurons of the hamster submandibular ganglion (SMG) have been studied using the whole-cell configuration of the patch-clamp technique. 2. Extracellular application of adenosine 5'-triphosphate (ATP) reversibly inhibited voltage-dependent Ca(2+) channel (VDCC) currents (I(Ca)) via G(i/o)-protein in a voltage-dependent manner. 3. Extracellular application of uridine 5'-triphosphate (UTP), 2-methylthioATP (2-MeSATP), alpha,beta-methylene ATP (alpha,beta-MeATP) and adenosine 5'-diphosphate (ADP) also inhibited I(Ca). The rank order of potency was ATP=UTP>ADP>2-MeSATP=alpha,beta-MeATP. 4. The P2 purinoceptor antagonists, suramin and pyridoxal-5-phosphate-6-azophenyl-2', 4'-disulfonic acid (PPADS), partially antagonized the ATP-induced inhibition of I(Ca), while coapplication of suramin and the P1 purinoceptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), virtually abolished I(Ca) inhibition. DPCPX alone partially antagonized I(Ca) inhibition. 5. Suramin antagonized the UTP-induced inhibition of I(Ca), while DPCPX had no effect. 6. Extracellular application of adenosine (ADO) also inhibited I(Ca) in a voltage-dependent manner via G(i/o)-protein activation. 7. Mainly N- and P/Q-type VDCCs were inhibited by both ATP and ADO via G(i/o)-protein betagamma subunits in seemingly convergence pathways.

The submandibular ganglion (SMG) neuron has been well established as the parasympathetic ganglion that innervates the submandibular and sublingual salivary glands. Thus this neuron plays a key role in salivary secretion. In a previous study, we reported that SMG possessed T-, L-, N-, P/Q- and R-type voltage-dependent calcium channels (VDCCs). In this study, we analyzed the contribution of the distinct subtypes of VDCCs currents (ICa) using the whole-cell configuration of the patch clamp technique in SMG neurons. In addition, we also investigated the effects of a strong voltage prepulse on the contributions of the subtypes of VDCCs. In SMG neuronal ICa without a prepulse, the mean percentages of L-, N-, P-, Q- and R-type were 39.7, 31.5, 10.6, 7.1 and 7.9%. In SMG neuronal ICa with prepulse, the mean percentages of L-, N-, P-, Q- and R-type were 37.2, 34.0, 14.0, 7.6 and 7.0%. Thus, these results showed that SMG possess multiple types of VDCCs and that N- and P-type VDCCs are facilitated by a prepulse in SMG neurons.

Angiotensin II (AngII) is one of the most important vasoconstrictive hormones but is also known to act as a neuromodulator and a neurotransmitter in the central and peripheral nervous systems. In a previous study, we have shown that AngII, mediated by AT(1) receptors, inhibits voltage-dependent calcium channel (VDCC) currents (I(Ca)) via G-proteins in submandibular ganglion (SMG) neurons. In this study, we further characterized the signal transduction of AngII-induced inhibition of I(Ca). Application of 1 microM AngII inhibited I(Ca) by 32.1+/-2.7% (mean+/-S.E.M., n=9). Intracellular dialysis of anti-G(q/11) antibodies attenuated these inhibition (8.8+/-1.3%, n=6). In addition, treatment of protein kinase C (PKC) activator and inhibitor also attenuated these inhibition (8.0+/-0.9 and 9.8+/-0.9%, n=6 and 9, respectively). We therefore conclude that AngII inhibits VDCC via G(q/11)-proteins involving in SMG neurons. In addition, such PKC-dependent pathways mediated mainly L-type VDCC inhibition.

In a previous study, we demonstrated that angiotensin II (AngII) inhibited voltage-dependent calcium channels (VDCCs) currents (ICa) in hamster submandibular ganglion (SMG) neurons. In sinoatrial node cells, it has been reported that AngII inhibits ICa by suppressing cyclic AMP production. In this study, to investigate the possible involvement of a cyclic AMP-cyclic AMP dependent protein kinase (PKA) pathway in the AngII-induced inhibition of ICa, effects of AngII were examined in SMG neurons after treatment with an activator and inhibitor of PKA. Neither pretreatment of neurons with membrane permeable cyclic AMP nor intracellular dialysis of PKA blocker attenuated the AngII-induced inhibition of ICa. These results indicate that AngII inhibited ICa via a cyclic AMP-PKA-independent mechanism in SMG neurons.

In this study, we investigated the effects of vasoactive intestinal polypeptide (VIP) and pituitary adenylate cyclase-activating polypeptide 1-38 (PACAP) on the voltage-gated calcium currents in hamster submandibular ganglion neurons. VIP and PACAP inhibited the high threshold voltage-gated calcium current in a voltage-independent and a concentration-dependent manner via the G protein-mediated pathway. L-, N- and P/Q-type components of the total maximum voltage-gated calcium current accounted for 48.0 +/- 3.1% (n = 4), 35.1 +/- 4.7% (n = 4), and 13.5 +/- 2.3% (n = 3) of the total peak amplitude, respectively. VIP at a concentration of 1 microM inhibited the L-type calcium current by 33.2% +/- 1.4% (n = 4), the N-type current by 31.0 +/- 3.6%, and the P/Q-type current by 3.2 +/- 1.1% (n = 3). PACAP at a concentration of 1 microM inhibited the L-type current by 35.6 +/- 5.7%, the N-type current by 34.4 +/- 3.1% (n = 4), and the P/Q-type current by 6.4 +/- 2.1% (n = 2). However, VIP and PACAP did not inhibit the low threshold voltage-gated (T-type) calcium current. The rank order of potency was PACAP > VIP. In experiments replacing GTP with GDP-beta-S, the inhibitory effects of VIP and PACAP were prevented. In experiments of double-pulse protocol, depolarizing conditioning pulses could not relieve the inhibition of total high threshold voltage-gated calcium currents produced by VIP and PACAP. Therefore, the inhibition of the high threshold voltage-gated calcium channels produced by VIP and PACAP in hamster parasympathetic neurons differed in its mechanisms from that of N-type calcium channels in rat sympathetic neurons.

We investigated the modulation of voltage dependent Ca(2+) currents by chemokine receptors in heterologous expression systems and neurons. Fractalkine, SDF-1alpha, RANTES and MDC inhibited the I(Ba) in CX3CR1-, CXCR4-, CCR5- and CCR4-expressing G1A1 cells, respectively. The I(Ba) inhibition was voltage-dependent, exhibited prepulse facilitation, and was blocked by N-ethylmaleimide and pertussis toxin pretreatment, indicating that it was mediated by Gi/Go. Some chemokines also inhibited the I(Ba) in subpopulations of dorsal root ganglion neurons and area postrema/nucleus tractus solitarius neurons. These data provide evidence that chemokines can potentially modulate neuronal signaling through the inhibition of neuronal Ca(2+) currents.

The calcium ion influx through voltage-dependent calcium channels (VDCCs) has a vital role in the control of neurotransmitter release and membrane excitability. Prepulse facilitation is a phenomenon in which a strong depolarizing pulse induces a form of the VDCCs that exhibits an increased opening probability in response to a given test potential; this persists for several seconds after repolarization. It has been reported that prepulse facilitation occurs via dissociation of the guanosine triphosphate (GTP)-binding proteins (G-proteins) from the VDCCs and that recovery from facilitation involves rebinding of the G-proteins. The heterotrimeric G-proteins act as switches that regulate information processing circuits connecting cell surface G-protein-coupled-receptors to a variety of effectors. In this study, we have studied the characterization of G-protein subtypes in prepulse facilitation of VDCCs currents (Ica) in hamster submandibular ganglion (SMG) neurons, using whole-cell patch clamp recordings. Under control conditions, with GTP (0.1 mM) in the recording pipette, the rate of prepulse facilitation was 19.0 +/- 1.9% (n = 13). Intracellular dialysis with GDP-beta-S (0.1 mM), G-protein blocker, and pretreatment of neurons with N-ethylmaleimide (NEM) (100 microM for 2 min), Gi/o blocker, attenuated the rate of prepulse facilitation. Intracellular dialysis of anti-Gq/11-antibody did not alter it. These results suggest that prepulse facilitation of VDCCs is due to Gi/o-types of G-protein, but not to the Gq/11-type, in SMG neurons.

Angiotensin II (Ang II) is one of the most important vasoconstrictive hormones but is also known to act as a neuromodulator and a neurotransmitter in the central and peripheral nervous systems. In a previous study, we have shown that Ang II, via AT1 receptors, induced depolarization by inhibition of M-type K(+) channels and SK channels in submandibular ganglion (SMG) neurons. In this study, we investigated the effects of Ang II on calcium channel current (I(Ca)) in acutely dissociated SMG neurons by the patch-clamp technique using the whole-cell configuration. Ang II inhibited total I(Ca) by 32.1+/-2.7%. The half-maximum inhibitory concentration (IC(50)) of Ang II for inhibiting I(Ca) was 0.8 microM. In the presence of 1 microM losartan, which is a selective antagonist of AT1 receptors, the effect of Ang II was attenuated (7.6+/-1.5%). Application of a strong depolarizing voltage prepulse did not affect the Ang II-induced inhibition of I(Ca) (32.8+/-2.8%). Intracellular dialysis of GDP-beta-S attenuated the inhibition of I(Ca) (6.8+/-2.1%). The mean percentage inhibitions of L-, N- and P/Q-type VDCCs by Ang II were 29.1+/-1.7, 16.3+/-6.0 and 1.2+/-0.8%, respectively, of the total I(Ca).

The calcium ion influx through voltage-dependent calcium channels (VDCCs) has a vital role in the control of neurotransmitter release and membrane excitability. The modulation of VDCCs controls the extent of calcium entry and thus provides a way of regulating neuronal function. Prepulse facilitation is a phenomenon in which a strong depolarizing pulse induces a form of the VDCCs that exhibits an increased opening probability in response to a given test potential that persists for several seconds after repolarization. In this study, we have studied the characterization of prepulse facilitation of VDCCs currents (Ica) in hamster submandibular ganglion (SMG) neurons, using whole-cell patch clamp recordings. In SMG neurons, application of a strong depolarizing prepulse caused a Ica. In 8 SMG neurons, rate of facilitation was 1.1 +/- 0.1. The greatest value of prepulse facilitation was obtained with prepulse to +100 mV, 10 ms duration in this neuron. The magnitude of facilitation was dependent on changing the interval between the -prepulse and the +prepulse and reached a maximum at a interval of 500 ms.

In this study, we investigated the modulation of calcium channels by vasoactive intestinal polypeptide (VIP) in hamster submandibular ganglion (SMG) cells using the whole-cell patch clamp technique. VIP (1 microM) inhibited the high voltage-gated calcium channels in the SMG neurons via activation of VIP receptors and G-protein coupled to these receptors, but did not affect the low voltage-gated calcium channels. VIP at 1 microM reduced the peak amplitude of the maximum high voltage-gated calcium current by 26.0 +/- 6.2% (mean +/- S.E., n = 5) and slowed the rising phase of the calcium current. The time to peak of the maximum high voltage-gated calcium current was prolonged from 16.8 msec to 22.4 msec. In a representative cell, 1 microM VIP reduced the peak amplitude of maximum high voltage-gated calcium current by 50.5% and the amplitude at the end of voltage step after 300 msec by 51.9%. The VIP-induced inhibition of the calcium current was produced in a voltage-dependent manner. Inhibition was maximal (50.5%) at the level of -20 mV. Therefore, VIP inhibits both transient and sustained types of high voltage-gated calcium channels in the hamster SMG neuron.

Both substance P (SP) and neurokinin A (NKA) are known as neurotransmitters of the submandibular ganglion (SMG) neurons. SP released from collaterals of the sensory nerves also regulates the excitability of SMG neurons. It has recently been shown that neurokinins (NK) inhibit calcium channels in various neurons. In this study, the effects of NK on voltage-dependent calcium channel current (I(Ca)) in SMG cells were investigated using the whole-cell patch-clamp recording method. NK-1 receptor agonist and SP caused inhibition of I(Ca) in SMG cells in a dose-dependent manner. NK-1 receptor agonist inhibited L-, N- and P/Q-type I(Ca) components. GDP-beta-S included in the pipette solution reduced the NK-1 receptor agonist-induced inhibition of I(Ca). In addition, NK-1 receptor agonist-induced inhibition of I(Ca) was reduced by stimulation of protein kinase C (PKC) but not cyclic AMP-dependent protein kinase (PKA). The results provided evidence for a signal transduction pathway in which calcium channel inhibition by NK receptors required activation of G-protein and PKC-affected step phosphorylation in SMG neurons.

It is well known that opioids produce inhibitory effects on neuronal activity and on synaptic transmission at most synapses. In this study, we have investigated the effects of opioids on the low voltage- and high voltage-activated calcium channels in acutely dissociated submandibular ganglion (SMG) neurons, using the whole-cell configuration of the patch-clamp technique. The kappa-opioid-receptor agonist U-50488H, the delta-opioid-receptor agonist [D-Pen 2,5]-enkephalin and the mu-opioid-receptor agonist [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin inhibited L-, N- and P/Q-type calcium-current components in a dose-dependent manner at 10 nM-1 microM, respectively, but not the T-type calcium current. These inhibitory effects were antagonized by naloxone (1 microM). The results showed that three types of opioid receptors regulate the L-, N- and P/Q-types of calcium channels, respectively, but not the T-type, in SMG neurones.