Abstract
Using immortalized hypothalamic GT1-7 neurons, which express the CB1 cannabinoid receptor (CB1R) and three Ca2+ channel types (T, R and L), we found that the CB1R agonist WIN 55,212-2 inhibited the voltage-gated Ca2+ currents by about 35%. The inhibition by WIN 55,212-2 (10 microM) was reversible and prevented by nifedipine (3 microM), suggesting a selective action on L-type Ca2+ channels (LTCCs). WIN 55,212-2 action exhibited all the features of voltage-independent Ca2+ channel modulation: (1) no changes of the activation kinetics, (2) equal depressive action at all potentials and (3) no facilitation following strong prepulses. At variance with WIN 55,212-2, the CB1R inverse agonist AM-251 (10 microM) caused 20% increase of Ca2+ currents. The inhibition of LTCCs by WIN 55,212-2 was prevented by overnight PTX-incubation and by intracellular perfusion with GDP-beta-S. The latter caused also a 20% Ca2+ current up-regulation. WIN 55,212-2 action was also prevented by application of the PKA-blocker H89 or by loading the neurons with 8-CPT-cAMP. Our results suggest that LTCCs in GT1-7 neurons are partially inhibited at rest due to a constitutive CB1R activity removed by AM-251 and GDP-beta-S. Activation of CB1R via PTX-sensitive G proteins and cAMP/PKA pathway selectively depresses LTCCs that critically control the synchronized spontaneous firing and pulsatile release of gonadotropin-releasing hormone in GT1-7 neurons.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Benzoxazines / pharmacology
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Calcium Channels, L-Type / physiology*
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Calcium Signaling / physiology*
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Cell Line
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Cyclic AMP / analogs & derivatives
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Cyclic AMP / metabolism*
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Cyclic AMP / pharmacology
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Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
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Cyclic AMP-Dependent Protein Kinases / metabolism*
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Down-Regulation
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Electrophysiology
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GTP-Binding Proteins / metabolism*
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Guanosine Diphosphate / analogs & derivatives
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Guanosine Diphosphate / pharmacology
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Hypothalamus / metabolism*
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Isoquinolines / pharmacology
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Mice
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Morpholines / pharmacology
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Naphthalenes / pharmacology
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Neuroendocrine Cells / metabolism*
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Piperidines / pharmacology
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Pyrazoles / pharmacology
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Receptor, Cannabinoid, CB1 / agonists
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Receptor, Cannabinoid, CB1 / antagonists & inhibitors
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Receptor, Cannabinoid, CB1 / metabolism*
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Signal Transduction / physiology
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Sulfonamides / pharmacology
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Thionucleotides / pharmacology
Substances
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Benzoxazines
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Calcium Channels, L-Type
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Isoquinolines
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Morpholines
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Naphthalenes
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Piperidines
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Pyrazoles
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Receptor, Cannabinoid, CB1
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Sulfonamides
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Thionucleotides
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Guanosine Diphosphate
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AM 251
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8-((4-chlorophenyl)thio)cyclic-3',5'-AMP
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(3R)-((2,3-dihydro-5-methyl-3-((4-morpholinyl)methyl)pyrrolo-(1,2,3-de)-1,4-benzoxazin-6-yl)(1-naphthalenyl))methanone
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guanosine 5'-O-(2-thiodiphosphate)
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Cyclic AMP
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Cyclic AMP-Dependent Protein Kinases
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GTP-Binding Proteins
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N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide