Identification and characterization of a novel synthetic cannabinoid CP 55,940 binder in rat brain cytosol |
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Authors: | Qureshi Jeffrey Saady Matt Cardounel Arturo Kalimi Mohammed |
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Institution: | (1) Department of Physiology, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia, USA |
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Abstract: | We have detected the presence of a specific 3H] CP 55,940 binder in the cytosol of rat cerebral cortex. Competition studies showed that only cold CP 55,940 and to a lesser extent D9THC was able to compete with 3H] CP 55,940; little competition was observed with either D8;THC or anandamide. Scatchard analysis of the data indicate the presence of two distinct binding components having affinity constants (Kd) of 0.97 ± 0.03 nM, 5.83 ± 0.08 nM, and Bmax of 3.31 ± 0.06 pmol/mg protein, 22.2 ± 1.2 pmol/mg protein respectively. The cytosolic CP 55,940 binder was heat stable up to 30øC. Besides the brain cytosol, lesser amounts of binding were also detected in the spleen, and testis. Liver, kidney and muscle cytosol preparations were found to be devoid of this binder. Unlike the previously characterized brain membrane cannabinoid receptor, this binder was found to be salt, sulfhydryl blocking reagents and nucleotide resistant. Interestingly, dithiothreitol (DTT), a protein-disulfide group reducing agent, inhibited the binding of 3H] CP-55,940 to the receptor and approximately 80% binding inhibition was obtained at a 5 mM concentration. Western blot analysis using anti-receptor antibody reveal the presence of a 95-110, 50 and 38 kDa band in the brain, spleen and testis cytosolic preparations. In conclusion, we have identified the presence of a novel CP 55,940 binder in rat cerebral cortex cytosol possessing biochemical properties distinct from those previously observed using rat cerebral cortex membrane cannabinoid receptor. |
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Keywords: | cannabinoid receptor rat brain cytosol CP 55 940 |
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