Identification of interleukin-1 receptors in mouse testis.
The cytokine interleukin-1 (IL-1) has been reported to alter reproductive functions through both effects in brain and direct actions at the level of the gonads. To further define the role of IL-1 at the gonads, we have used 125I-labeled recombinant human IL-1 to identify and characterize IL-1 receptors in crude membrane preparations of mouse (C57BL/6) testis and to study the distribution of IL-1-binding sites using autoradiography. In preliminary homogenate binding and autoradiographic studies, [125I]IL-1 alpha showed significantly higher specific binding than [125I]IL-1 beta. Thus, [125I]IL-1 alpha was used in all subsequent assays. The binding of [125I]IL-1 alpha was linear over a broad range of membrane protein concentrations, saturable, and reversible, and on Scatchard analysis revealed an equilibrium dissociation constant (Kd) of 82 +/- 4 pM and a maximum number of binding sites (Bmax) of 10.8 +/- 1.5 fmol/mg protein. In competition studies, IL-1 alpha, IL-1 beta, and a weak IL-1 beta analog IL-1 beta+ inhibited [125I]IL-1 alpha binding to mouse testis in parallel with their relative bioactivities in immune assays, with inhibitory binding affinity constant (Ki) values of 14.2 +/- 1.7, 88.8 +/- 5.7, and 7183.3 +/- 603 pM, respectively; rat/human CRF and human tumor necrosis factor showed no effect on [125I]IL-1 alpha binding. In autoradiographic studies, IL-1 receptors were heterogeneously distributed, with highest densities present in the luminal border of the epididymis and interstitial areas of the testis. After hypophysectomy, the testes were significantly atrophied, and homogenate binding and autoradiographic studies showed that while the total number of binding sites per testis was significantly decreased in hypophysectomized mice in proportion to the reduction in testicular mass, there was no apparent change in the relative density of IL-1 receptors. These data provide the first identification of IL-1 receptors in testis and provide further support for a physiological role for IL-1 to alter reproductive functions through a direct effect at the gonads.
Takao
T
T
Neurobiology Laboratory, National Institute on Drug Abuse, Addiction Research Center, Baltimore, Maryland 21224.
Mitchell
W M
WM
Tracey
D E
DE
De Souza
E B
EB
D000818:Animals; D001345:Autoradiography; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D003346:Corticotropin-Releasing Hormone / Q000494:pharmacology; D006168:Guinea Pigs; D007016:Hypophysectomy; D007375:Interleukin-1 / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D051381:Rats; D011971:Receptors, Immunologic / Q000032:analysis* / Q000378:metabolism; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins / Q000378:metabolism; D013737:Testis / Q000032:analysis* / Q000378:metabolism; D014018:Tissue Distribution; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D007375:Interleukin-1; D011971:Receptors, Immunologic; D017472:Receptors, Interleukin-1; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D003346:Corticotropin-Releasing Hormone