[125I]-labeled forskolin analogs which discriminate adenylyl cyclase and a glucose transporter: pharmacological characterization and localization of binding sites in rat brain by in vitro receptor autoradiography

NM Appel, JD Robbins, EB De Souza and KB Seamon

Division of Research and Testing HFD-472, Food and Drug Administration, Laurel, Maryland.

Aminoalkylcarbamate derivatives of forskolin have been synthesized at the 6- and 7-hydroxyl positions which have different selectivity for adenylyl cyclase and a glucose transporter, respectively. They were radioiodinated using the Bolton-Hunter reagent to yield [125I]-2-[3-(4- hydroxy-3-iodophenyl)propanamido]-N-ethyl-6- (aminocarbonyl)forskolin ([125I]6-IHPP-Fsk) and [125I]-2-[3-(4-hydroxy-3- iodophenyl)(propanamidol]-N-ethyl-7- (aminocarbonyl)-7- desacetylforskolin ([125I]7-IHPP-Fsk) and tested as autoradiographic probes for adenylyl cyclase and a glucose transporter. In slide-mounted rat brain sections [125I]6-IHPP-Fsk binding was potently inhibited by 1 microM 6-HPP-Fsk (95%) but unaffected by 500 mM D-glucose. In contrast, [125I]7-IHPP-Fsk was only partially inhibited by 1 microM 6-HPP-Fsk (37%), but residual [125I]7-IHPP-Fsk binding was further inhibited 56% by 500 mM D-glucose. These data suggest that while [125I]6-IHPP-Fsk binds exclusively to adenylyl cyclase, a significant fraction of [125I]7-IHPP-Fsk is binding to a glucose transporter in brain. Autoradiographic patterns of [125I]6-IHPP-Fsk and glucose-sensitive [125I]7-IHPP-Fsk binding were different. [125I]6-IHPP-Fsk binding was heterogeneously distributed and resembled [3H] forskolin binding. Highest densities of binding sites were noted in olfactory tubercle, caudate putamen, nucleus accumbens, pyramidal and granule cell layers of hippocampus, molecular layer of cerebellum and substantia nigra. In contrast, of glucose-sensitive [125I]7-IHPP-Fsk, binding appeared more homogeneous and similar to [3H]cytochalasin B, a compound which inhibits glucose transport. Highest densities of binding were noted in caudate putamen, nucleus accumbens, cerebral cortex and molecular layer of cerebellum.(ABSTRACT TRUNCATED AT 250 WORDS)

Volume 263, Issue 3, pp. 1415-1423, 12/01/1992
Copyright © 1992 by American Society for Pharmacology and Experimental Therapeutics