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3′,5′‐Cyclic adenosine monophosphate as an intracellular second messenger of luteinizing hormone: Application of the forskolin criteria
Author(s) -
Adashi Eli Y.,
Resnick Carol E.
Publication year - 1986
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.240310304
Subject(s) - forskolin , medicine , endocrinology , luteinizing hormone , adenosine , phosphodiesterase , gonadotropin , second messenger system , human chorionic gonadotropin , testosterone (patch) , androgen , cyclic nucleotide , cyclic adenosine monophosphate , phosphodiesterase inhibitor , stimulation , chemistry , extracellular , biology , cyclase , hormone , enzyme , nucleotide , biochemistry , receptor , gene
The role of adenosine 3′,5′‐cyclic monophosphate (cAMP) as an intracellular second messenger of luteinizing hormone (LH) was reinvestigated in vitro with diterpene forskolin, a highly specific activator of adenylate cyclase. Treatment of cultured testicular cells from adult hypophysectomized rats with increasing concentrations (10 −7 –10 −4 M) of forskolin produced dose‐dependent increments in cAMP and testosterone accumulation. Concomitant blockade of cAMP‐phosphodiesterase activity with 3‐isobutyl‐1‐methyl‐xanthine (10 −4 M) resulted in significant (P < 0.05) enhancement of the forskolin effect for all but the 10 −4 M forskolin dose. Potency evaluation as judged by half‐maximal stimulation of testosterone accumulation revealed median effective doses (mean ± SE) of 1.25 ± 0.2 × 10 −5 , 1.7 ± 0.5 × 10 −5 , and 2.5 ± 0.4 × 10 −10 M for forskolin, N 6 , O 2 ′‐dibutyryl cAMP (Bt 2 cAMP), and human chorionic gonadotropin (hCG), respectively. Examination of the time requirements of forskolin disclosed time‐dependent increments in the accumulation of extracellular cAMP and testosterone, the earliest significant (P < 0.05) increases being noted by 6 hr of treatment. In comparison, a minimal time requirement of ⩽ 12hr was noted for hCG‐ and choleragen‐stimulated androgen biosynthesis, whereas the apparent onset of action of Bt 2 cAMP was delayed to the 24‐hr time point. Although 10 −7 M of forskolin by itself did not alter the accumulation of testosterone, its addition resulted in substantial amplification of the hCG effect, producing a 4.6‐fold reduction in the median effective dose (ED 50 ) of hCG. Moreover, concurrent treatment with this functionally inert dose of forskolin rendered steroidogenically inert doses of hCG (eg, 10 −11 or 3 × 10 −11 M) steroidogenically potent. However, combined treatment with maximally stimulatory doses of Bt 2 cAMP (10 −4 M) and one of several testicular cell agonists [forskolin (10 −4 M), choleragen (10 −9 M) or hCG (10 −9 M)] did not prove additive. Taken together, our findings indicate that forskolin, like LH, is capable of stimulating testicular cAMP generation as well as androgen biosynthesis and that a functionally inert low dose of forskolin can significantly amplify LH hormonal action. Inasmuch as forskolin‐stimulated and forskolin‐amplified hormonal action are acceptable as novel criteria of cAMP dependence, our observations provide new evidence in keeping with the notion that cAMP may be in intracellular second messenger of LH.