Pancreatic polypeptide inhibits calcium channels in rat sympathetic neurons via two signaling pathways

1995 ◽  
Vol 73 (3) ◽  
pp. 1323-1328 ◽  
Author(s):  
L. P. Wollmuth ◽  
M. S. Shapiro ◽  
B. Hille
Keyword(s):  
G Protein ◽  
Pertussis Toxin ◽  
Pancreatic Polypeptide ◽  
Sympathetic Neurons ◽  
Whole Cell ◽  
Adult Rat ◽  
Voltage Dependent ◽  
Cervical Ganglion ◽  
Gamma S ◽  
Ca2 Channels

1. We studied modulation of N-type Ca2+ channels in adult rat superior cervical ganglion (SCG) neurons by pancreatic polypeptide (PP) using whole cell clamp. In large (> 20 pF) SCG neurons, PP inhibited ICa (35 +/- 2%, mean +/- SE) in a concentration-dependent fashion, with one-half maximal inhibition at 19 nM. 2. One-third of the inhibition was blocked by pertussis toxin, about one-half was blocked by N-ethylmaleimide (NEM) treatments, and about one-half was voltage dependent. The NEM-insensitive component of the PP inhibition was voltage independent and not significantly blocked by intracellular Ca2+ chelators. 3. The NEM-insensitive component was only weakly attenuated by GDP-beta-S, and moderately reversible with guanosine 5'-triphosphate (GTP)-gamma-S, in the whole cell pipette, leaving open the possibility that it is not mediated by a G protein. 4. Hence, PP inhibits ICa via two mechanisms: one G-protein-mediated and the other possibly G-protein independent. The former pathway is sensitive to pertussis toxin (PTX) and NEM, voltage dependent, and shared by several other transmitters in these cells. The latter pathway is PTX-and NEM-insensitive, not voltage dependent, and not affected by the presence of intracellular Ca2+ chelators.

Adenosine modulates voltage-gated Ca2+ channels in adult rat sympathetic neurons

1993 ◽  
Vol 70 (2) ◽  
pp. 610-620 ◽  
Author(s):  
Y. Zhu ◽  
S. R. Ikeda
Keyword(s):  
G Protein ◽  
Pertussis Toxin ◽  
Adenosine Receptor ◽  
Sympathetic Neurons ◽  
Current Component ◽  
Tail Current ◽  
Adult Rat ◽  
Selective Agonist ◽  
Voltage Dependent ◽  
Ca2 Channels

1. Ca(2+)-channel modulation by adenosine was investigated in enzymatically dispersed adult rat superior cervical ganglion (SCG) neurons using the whole-cell variant of the patch-clamp technique. 2. Adenosine produced a concentration-dependent decrease in the Ca(2+)-current amplitude with an EC50 of 174 nM and maximum inhibition of 36%. The effects of adenosine on the Ca2+ current were both time and voltage dependent. The inhibition was maximal at +10 mV and decreased at either hyperpolarizing or depolarizing potentials. 3. The inhibitory response desensitized after prolonged (> 1 min) exposure to 10 microM adenosine, whereas multiple brief (< 30 s) applications slightly decreased the subsequent response. 4. Adenosine-induced Ca2+ current inhibition was mediated by an A1-type adenosine receptor, because the half-maximal inhibition value for an A1 receptor selective agonist, chloro-N-cyclopentyladenosine, was 1,000-fold lower than that for an A2 receptor selective agonist, 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarbozamido adenosine hydrochloride (33 nM vs. 40 microM, respectively). 5. A guanine nucleotide binding protein (G protein) appeared to be involved in the action of adenosine, because: 1) the adenosine-induced current inhibition could be largely relieved by depolarizing voltage prepulses; 2) tail current analysis revealed that adenosine shifted Ca(2+)-channel activation to more depolarized potentials; and 3) adenosine inhibition was abolished by 2 mM intracellular guanosine 5'-O-(2-thiodiphosphate) or 500 ng/ml pertussis toxin pretreatment. 6. Adenosine did not appear to inhibit L-type Ca2+ channels, because the prolonged tail current component induced by the dihydropyridine "agonist" 2,6-dimethy-3-carbomethoxy-5-nitro-4-(2-trifluoromethyl-phenyl)- 1,4-dihydropyridine (2 microM) was not affected by adenosine. 7. Adenosine-induced inhibition was reduced to approximately 15% after application of 10 microM omega-conotoxin GVIA, suggesting that adenosine primarily inhibits N-type Ca2+ channels. The Ca(2+)-current component resistant to omega-conotoxin GVIA was also resistant to omega-agatoxin IVA (200 nM), suggesting a lack of P-type of Ca2+ channels in SCG neurons. 8. In conclusion, adenosine produces a dose-, time-, and voltage-dependent inhibition of Ca2+ currents in SCG neurons. Adenosine acts on an A1 adenosine receptor subtype in SCG neurons via a pertussis toxin-sensitive G protein to inhibit N-type Ca2+ channels and an unidentified Ca(2+)-current component. Modulation of Ca2+ currents by adenosine may be an important mechanism for its inhibitory effect on neurotransmitter release in sympathetic neurons.

Modulation of Ca(2+)-channel currents by protein kinase C in adult rat sympathetic neurons

1994 ◽  
Vol 72 (4) ◽  
pp. 1549-1560 ◽  
Author(s):  
Y. Zhu ◽  
S. R. Ikeda
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
G Protein ◽  
G Proteins ◽  
Sympathetic Neurons ◽  
Adult Rat ◽  
Kinase C ◽  
Channel Inhibition ◽  
Channel Currents ◽  
Ca2 Channels

1. Modulation of Ca(2+)-channel currents by phorbol-12-myristate-13-acetate (PMA) was investigated in acutely dissociated adult rat superior cervical ganglion neurons using the whole cell variant of the patch-clamp technique. 2. PMA (500 nM) increased the current amplitudes, accelerated the inactivation of step currents, retarded the deactivation of tail currents, and shifted the tail current activation to more negative potentials. 3. The effects of PMA were concentration and voltage dependent and mediated through activation of protein kinase C (PKC). PMA also increased Ca2+ currents recorded with the perforated patch technique. 4. PMA affected the N-type Ca2+ channels and an omega-conotoxin GVIA-resistant current component. Ca2+ currents affected by PMA were not sensitive to omega-agatoxin IVA or nimodipine. 5. PMA not only attenuated Ca(2+)-channel inhibition induced by alpha 2-adrenoceptor agonist, which modulates Ca2+ channels via a pertussis toxin (PTX)-sensitive pathway, but also attenuated current inhibition by vasoactive intestinal polypeptide, which modulates Ca2+ channels via a PTX-insensitive but cholera toxin-sensitive pathway. 6. PMA reversed Ca(2+)-channel inhibition induced by tonic activation of G-protein in the absence of neurotransmitter (even in neurons pretreated with PTX) or induced by activation of G-proteins with guanosine 5'-O-(3-thiotriphosphate) (GTP)-gamma-S. 7. Inhibition of phosphatase by okadaic acid or substitution of Ba2+ for Ca2+ in the external solutions accelerated the PMA effect. 8. Our results suggest that activation of PKC antagonizes G-protein mediated inhibition of Ca2+ channels by shifting Ca2+ channels from the “reluctant” state to the “willing” state. The G-proteins and, more likely, the N-type Ca2+ channels may be the target of PKC phosphorylation. Protein phosphatases may be involved in counteracting the PKC phosphorylation in rat sympathetic neurons.

scholarly journals N-Arachidonoyl l-Serine, a Putative Endocannabinoid, Alters the Activation of N-Type Ca2+ Channels in Sympathetic Neurons

2008 ◽  
Vol 100 (2) ◽  
pp. 1147-1151 ◽  
Author(s):  
Juan Guo ◽  
Damian J. Williams ◽  
Stephen R. Ikeda
Keyword(s):  
G Protein ◽  
Sympathetic Neurons ◽  
G Protein Coupled Receptors ◽  
Whole Cell Patch Clamp ◽  
Voltage Dependent ◽  
Nervous System Function ◽  
Ganglion Neurons ◽  
G Protein Coupled ◽  
Hyperpolarizing Shift ◽  
Ca2 Channels

The effect of N-arachidonoyl l-serine (ARA-S), a recently discovered lipoamino acid found in the CNS, on N-type Ca2+ channels of rat sympathetic ganglion neurons was determined using whole cell patch clamp. Application of ARA-S produced a rapid and reversible augmentation of Ca2+ current that was voltage dependent and resulted from a hyperpolarizing shift in the activation curve. ARA-S did not influence G protein modulation of Ca2+ channels and appeared to act independently of G-protein-coupled receptors. These findings provide a foundation for investigating possible roles for ARA-S in nervous system function.

scholarly journals G protein control of potassium channel activity in a mast cell line.

1990 ◽  
Vol 95 (2) ◽  
pp. 205-227 ◽  
Author(s):  
M A McCloskey ◽  
M D Cahalan
Keyword(s):  
Cell Line ◽  
G Protein ◽  
Pertussis Toxin ◽  
Inward Rectifier ◽  
K Channel ◽  
Half Time ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
K Channels ◽  
Gamma S

Using the patch-clamp technique, we studied regulation of potassium channels by G protein activators in the histamine-secreting rat basophilic leukemia (RBL-2H3) cell line. These cells normally express inward rectifier K+ channels, with a macroscopic whole-cell conductance in normal Ringer ranging from 1 to 16 nS/cell. This conductance is stabilized by including ATP or GTP in the pipette solution. Intracellular dialysis with any of three different activators of G proteins (GTP gamma S, GppNHp, or AlF-4) completely inhibited the inward rectifier K+ conductance with a half-time for decline averaging approximately 300 s after "break-in" to achieve whole-cell recording. In addition, with a half-time averaging approximately 200 s, G protein activators induced the appearance of a novel time-independent outwardly rectifying K+ conductance, which reached a maximum of 1-14 nS. The induced K+ channels are distinct from inward rectifier channels, having a smaller single-channel conductance of approximately 8 pS in symmetrical 160 mM K+, and being more sensitive to block by quinidine, but less sensitive to block by Ba2+. The induced K+ channels were also highly permeable to Rb+ but not to Na+ or Cs+. The current was not activated by the second messengers Ca2+, inositol 1,4,5-trisphosphate, inositol 1,3,4,5-tetrakisphosphate, or by cyclic AMP-dependent phosphorylation. Pretreatment of cells with pertussis toxin (0.1 microgram/ml for 12-13 h) prevented this current's induction both by guanine nucleotides and aluminum fluoride, but had no effect on the decrease in inward rectifier conductance. Since GTP gamma S is known to stimulate secretion from patch-clamped rat peritoneal mast cells, it is conceivable that K+ channels become inserted into the plasma membrane from secretory granules. However, total membrane capacitance remained nearly constant during appearance of the K+ channels, suggesting that secretion induced by GTP gamma S was minimal. Furthermore, pertussis toxin had no effect on secretion triggered by antigen, and triggering of secretion before electrical recording failed to induce the outward K+ current. Finally, GTP gamma S activated the K+ channel in excised inside-out patches of membrane. We conclude that two different GTP-binding proteins differentially regulate two subsets of K+ channels, causing the inward rectifier to close and a novel K+ channel to open when activated.

The neuropeptide FMRFa both inhibits and enhances the Ca2+ current in dissociated Helix neurons via independent mechanisms

1991 ◽  
Vol 65 (6) ◽  
pp. 1517-1527 ◽  
Author(s):  
J. L. Yakel
Keyword(s):  
G Protein ◽  
Cell Voltage ◽  
Inhibitory Response ◽  
Central Neurons ◽  
S 100 ◽  
Helix Neurons ◽  
20 Nm ◽  
Gamma S ◽  
Ca2 Channels ◽  
Rate Of Activation

1. The modulation of the voltage-activated Ca2+ current by the neuropeptide Phe-Met-Arg-Phe-NH2 (FMRFa) was investigated in dissociated central neurons from Helix aspersa using whole-cell voltage-clamp recording techniques. External Ba2+ was always used as the charge carrier in this study, and the intracellular Ca2+ concentration was buffered to 20 nM with ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA). 2. Run-down of the Ca2+ currents was not a problem as long as the neurons were dialyzed with a patch electrode filling solution containing ATP (1 or 2 mM). In ATP-dialyzed neurons, the rate of inactivation of the calcium current increased with time without any significant change in the rate of activation. However, when neurons were dialyzed with guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S; 100 microM; with ATP), the rate of inactivation decreased with time. There was no effect of GTP gamma S on the rate of activation of the Ca2+ current. This suggests that guanosine 5'-triphosphate (GTP)-binding proteins (G proteins) are able to modulate the rate of inactivation of the Ca2+ current in Helix neurons. 3. FMRFa both decreased and enhanced the amplitude of the Ca2+ current in these neurons. This inhibition was observed in most neurons, while the enhancement was observed in 20% of the neurons. Although the enhancement usually was preceded by the inhibitory response, sometimes the enhancement was observed separately. 4. The FMRFa-induced inhibition of the Ca2+ current usually consisted of a decrease in both the amplitude and the rate of inactivation of this current, effects that were reduced as the membrane potential was stepped to more depolarized potentials. A pertussis toxin (PTX)-sensitive G protein mediated this response, whereas no evidence was found to suggest the involvement of any known intracellular messenger. Therefore this inhibition may have resulted from a direct coupling between the FMRFa receptor and the Ca2+ channels via a PTX-sensitive G protein. 5. Arachidonic acid (100 microM) irreversibly reduced the amplitude of the Ca2+ current, but it did not alter the relative inhibition of this current by FMRFa. 6. The FMRFa-induced enhancement of the Ca2+ current was difficult to study because it was observed infrequently, and was rarely observed independently of the FMRFa-induced inhibitory response. In addition, the ability of FMRFa to enhance this current usually disappeared with time.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Intracellular Na+inhibits voltage-dependent N-type Ca2+channels by a G protein βγ subunit-dependent mechanism

2004 ◽  
Vol 556 (1) ◽  
pp. 121-134 ◽  
Author(s):  
Yakov Blumenstein ◽  
Olexandr P. Maximyuk ◽  
Natalia Lozovaya ◽  
Natalia M. Yatsenko ◽  
Nataly Kanevsky ◽  
...  
Keyword(s):  
G Protein ◽  
Voltage Dependent ◽  
Ca2 Channels ◽  
Dependent Mechanism

Norepinephrine Inhibits a Toxin Resistant Ca2+ Current in Carotid Body Glomus Cells: Evidence for a Direct G Protein Mechanism

1999 ◽  
Vol 81 (1) ◽  
pp. 225-233 ◽  
Author(s):  
Jeffrey L. Overholt ◽  
Nanduri R. Prabhakar
Keyword(s):  
Receptor Antagonist ◽  
G Protein ◽  
G Proteins ◽  
Carotid Body ◽  
Adrenergic Receptor ◽  
Subunit Interaction ◽  
Whole Cell ◽  
Glomus Cells ◽  
Voltage Dependent ◽  
Carotid Body Glomus Cells

Overholt, Jeffrey L. and Nanduri R. Prabhakar. Norepinephrine inhibits a toxin resistant Ca2+ current in carotid body glomus cells: evidence for a direct G protein mechanism. J. Neurophysiol. 81: 225–233, 1999. Previous studies have demonstrated that endogenous norepinephrine (NE) inhibits carotid body (CB) sensory discharge, and the cellular actions of NE have been associated with inhibition of Ca2+ current in glomus cells. The purpose of the present study was to elucidate the characteristics and mechanism of NE inhibition of whole cell Ca2+ current isolated from rabbit CB glomus cells and to determine the type(s) of Ca2+ channel involved. NE (10 μM) inhibited 24 ± 2% (SE) of the macroscopic Ca2+ current measured at the end of a 25 ms pulse to 0 mV and slowed activation of the current. The α2 adrenergic receptor antagonist, SK&F 86466, attenuated these effects. Inhibition by NE was fast and voltage-dependent i.e., maximal at −10 mV and then diminished with stronger depolarizations. This is characteristic of G protein βγ subunit interaction with the α1 subunit of certain Ca2+ channels, which can be relieved by depolarizing steps. A depolarizing step (30 ms to +80 mV) significantly increased (14 ± 1%) current in the presence of NE, whereas it had no effect before application of NE (1 ± 1%). To further test for the involvement of G proteins, NE was applied to cells where intracellular GTP was replaced by GDP-βS. NE had little or no effect on Ca2+ current in cells dialyzed with GDP-βS. To determine whether NE was inhibiting N- and/or P/Q-type channels, we applied NE in the presence of ω-conotoxin MVIIC (MVIIC). In the presence of 2.5 μM MVIIC, NE was equally potent at inhibiting the Ca2+ current (23 ± 4% vs. 23 ± 4% in control), suggesting that NE was not exclusively inhibiting N- or P/Q-type channels. NE was also equally potent (30 ± 2% vs. 26 ± 4% in control) at inhibiting the Ca2+ current in the presence of 2 μM nisoldipine, suggesting that NE was not inhibiting L-type channels. Further, NE inhibited a significantly larger proportion (47 ± 6%) of the resistant Ca2+ current remaining in the presence of NISO and MVIIC. These results suggest that NE inhibition of Ca2+ current in rabbit CB glomus cells is mediated in most part by effects on the resistant, non L-, N-, or P/Q-type channel and involves a direct G protein βγ interaction with this channel.

Voltage-dependent facilitation of calcium channels in rat neostriatal neurons

1996 ◽  
Vol 76 (4) ◽  
pp. 2290-2306 ◽  
Author(s):  
W. J. Song ◽  
D. J. Surmeier
Keyword(s):  
Polymerase Chain Reaction Amplification ◽  
Modal Shift ◽  
Whole Cell ◽  
Adult Rat ◽  
Class D ◽  
Current Voltage ◽  
Cell Recording ◽  
Voltage Dependent ◽  
Reaction Amplification ◽  
Class C

1. Voltage-dependent facilitation of Ca2+ channels was studied in acutely isolated adult rat neostriatal neurons. Particular attention was paid to the facilitation of L-type channels. 2. In the absence of neuromodulators, the current-voltage relationship for whole cell Ba2+ currents was enhanced by a prepulse to +100 mV. The median enhancement at -20 mV was nearly 60%. The voltage dependence and kinetics of the processes underlying the facilitation were similar to those reported in other neurons. N-, P-, Q-, and L-type currents contributed to the observed facilitation. 3. Voltage-dependent facilitation of L-type currents was studied by subtracting nifedipine-insensitive currents from control currents. Although the kinetics were similar to those of the whole cell currents, the half-activation voltage for facilitation of L-type currents [half-activation voltage (Vh) = -0.6 mV, slope factors (Vc) = 11.8 mV, [n = 5] was significantly less depolarized than that of the pooled currents (Vh = 47.3 mV, Vc = 12.3 mV, n = 7). 4. Repetitive depolarization with spikelike waveforms was also able to induce facilitation of L-type currents, suggesting that facilitation was not simply a consequence of a modal shift in gating like that induced by Bay K 8644. 6. Combined whole cell recording and single-cell reverse transcription-polymerase chain reaction amplification revealed that neostriatal medium spiny neurons expressed detectable levels of either class C or class D L-type channel alpha 1, subunit mRNA. Both neurons expressing class C L-type channels and neurons expressing class D L-type channels exhibited voltage-dependent facilitation.

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