Interaction between calcium channel ligands and guanine nucleotides in cultured rat sensory and sympathetic neurones.
1. Voltage-activated Ca2+ channel currents were recorded from cultured rat dorsal root ganglion (DRG) neurones using the whole-cell clamp technique with Ba2+ as the charge carrier. 2. Inclusion of the GTP analogue guanosine 5'-O-3-thiotriphosphate (GTP-gamma-S, 500 microM) or guanylylimidodiphosphate (GMP-PNP, 500 microM) or GTP itself (1 mM) in the patch pipette solution resulted in a smaller, slowly activating Ca2+ channel current which did not inactivate during a 100 ms voltage step. This current was inhibited by CdCl2 (10-100 microM) and omega-conotoxin (1 microM). 3. Nifedipine (5 microM), (-)-(R)-201-791 (5 microM), D600 (10 microM), and diltiazem (30 microM) inhibited Ca2+ channel currents recorded from control neurones, although in some cells a biphasic response was observed, with an initial increase preceding the inhibition of the currents. In the presence of internal GTP-gamma-S, at a holding potential (VH) of -80 mV, only potentiation of the Ca2+ channel current was observed in the presence of all three Ca2+ channel ligands. Internal GMP-PNP, while less effective than GTP-gamma-S, also resulted in D600 showing an agonist response. Similarly, in the presence of internal GTP (1 mM), (-)-(R)-202-791 gave a prolonged agonist response. 4. Nifedipine, whether acting as an antagonist in control cells or as an agonist in GTP-gamma-S-containing cells, induced a shift to more hyperpolarized potentials of the steady-state inactivation curves. 5. Potentiation of Ca2+ channel currents induced by D600 in GTP-gamma-S-containing cells, was not observed when the neurones were pre-treated with pertussis toxin. The presence of internal GDP-beta-S (500 microM) did not significantly alter the maximum inhibitory action of D600 compared with controls. However, 1 mM-GDP-beta-S increased the rate of onset of inhibition by (-)-(R)-202-791. 6. Depolarizing VH to -30 mV accelerated the onset of inhibition induced by the Ca2+ channel ligands in control cells. In the presence of internal GTP-gamma-S at VH -30 mV, biphasic responses were produced by all the Ca2+ channel antagonist ligands with initial stimulation for 1-2 min being followed by inhibition of the Ca2+ channel currents. 7. The agonist actions of (+)-(S)-202-791 were potentiated by the presence of internal GTP-gamma-S. 8. The expression of an agonist response to (-)-(R)-202-791 induced by internal GTP-gamma-S was also present in sympathetic neurones cultured from adult rat superior cervical ganglion (SCG).(ABSTRACT TRUNCATED AT 400 WORDS)
Dolphin
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Department of Pharmacology, St George's Hospital Medical School, London.
Scott
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D000818:Animals; D002120:Calcium Channel Agonists / Q000494:pharmacology*; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D004095:Dihydropyridines / Q000494:pharmacology; D005711:Gallopamil / Q000494:pharmacology; D005727:Ganglia, Spinal / Q000166:cytology* / Q000187:drug effects; D005728:Ganglia, Sympathetic / Q000166:cytology* / Q000187:drug effects; D006150:Guanine Nucleotides / Q000494:pharmacology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D008024:Ligands; D009475:Neurons, Afferent / Q000187:drug effects / Q000502:physiology*; D009543:Nifedipine / Q000494:pharmacology; D051381:Rats; D013873:Thionucleotides / Q000494:pharmacology
D002120:Calcium Channel Agonists; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004095:Dihydropyridines; D006150:Guanine Nucleotides; D008024:Ligands; D013873:Thionucleotides; D006153:Guanosine Diphosphate; D006165:Guanylyl Imidodiphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D005711:Gallopamil; C023427:guanosine 5'-O-(2-thiodiphosphate); D006160:Guanosine Triphosphate; C045796:SAN 202791; D009543:Nifedipine