title: Inositol lipid-mediated signalling in response to endothelin and ATP in the mammalian testis.
abstract: The testis is a complex organ in which local control is achieved by signalling between its constituent cells. Herein we describe the responses of cultured rat testicular cells and a mouse Sertoli cell-line to stimulation by endothelin and ATP, and elsewhere we have shown that rat peritubular myoid cells possess phosphoinositidase C-coupled V1a-vasopressin receptors identical to those of liver (Howl, J. et al, 1995, Endocrinology 136: 2206-2213). 1. Peritubular myoid cells from pre-pubertal rats responded through ETA receptors with PtdIns(4,5)P2 hydrolysis [EC50 for endothelin-1 (ET-1) approximately 0.4 nM], elevation of intracellular [Ca2+], and tyrosine phosphorylation of a variety of cellular proteins. They also showed enhanced adenylate cyclase activity, with an EC50 for ET-1 of approximately 3 nM, also through ETA receptors. Pharmacological elevation of [cAMP] did not immediately change the ET-1-stimulated formation of inositol phosphates, but attenuated the response after several hours. 2. Pre-pubertal rat Sertoli cells showed no detectable responses to ET-1, but responded to FSH with elevated [cAMP] and to ATP with PtdIns(4,5)P2 hydrolysis. PtdIns(4,5)P2 hydrolysis was equally responsive to ATP and UTP, and so appears to be activated by P2U-purinergic receptors. This response was enhanced by protein kinase C inhibition and attenuated by PKC activation. 3. Despite its lack of effect on rat Sertoli cells in primary culture, ET-1 provoked PtdIns(4,5)P2 hydrolysis in the TM4 murine Sertoli cell line (EC50 approximately 0.6 nM), and this response was negatively regulated by protein kinase C activation. 5. No receptor-stimulated activation of phosphoinositase C was detected in 'germ cell' populations, but the non-specific G protein activator A1F4-provoked inositol phosphate accumulation in these cells, so demonstrating their potential to respond through yet to be identified G protein-coupled receptors with phosphoinositidase C activation. 6. Immunoblotting studies showed the presence in rat testis of phosphoinositidase C-beta 1 and the alpha-subunits(s) of the G-protein(s) Gq and/or G11. These studies show that testicular myoid and Sertoli cells use at least three G protein-coupled receptors (V1a-vasopressins, ETA-endothelin and P2U-purinergic) to signal through phosphoinositidase C activation, that ET-1 can activate multiple signalling pathways in myoid cells, and that the ET-1-stimulated phosphoinositidase C responses of myoid and Sertoli cells have different regulatory characteristics.
authors.affiliation: Centre for Clinical Research in Immunology and Signalling, University of Birmingham, UK.
authors.forename: S A
authors.initials: SA
authors.lastname: Rudge
authors.forename: P J
authors.initials: PJ
authors.lastname: Hughes
authors.forename: G R
authors.initials: GR
authors.lastname: Brown
authors.forename: R H
authors.initials: RH
authors.lastname: Michell
authors.forename: C J
authors.initials: CJ
authors.lastname: Kirk
mesh_terms: D000255:Adenosine Triphosphate / Q000502:physiology; D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D016232:Endothelins / Q000502:physiology*; D004789:Enzyme Activation; D005640:Follicle Stimulating Hormone / Q000502:physiology; D019204:GTP-Binding Proteins / Q000502:physiology; D007295:Inositol Phosphates / Q000502:physiology; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D010716:Phosphatidylinositols / Q000502:physiology; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D019000:Phosphotyrosine / Q000378:metabolism; D011493:Protein Kinase C / Q000502:physiology; D051381:Rats; D017466:Receptors, Endothelin / Q000502:physiology*; D011983:Receptors, Purinergic / Q000502:physiology*; D012708:Sertoli Cells / Q000502:physiology*; D012741:Sexual Maturation; D015398:Signal Transduction; D013737:Testis / Q000502:physiology*
chemical_list: D016232:Endothelins; D007295:Inositol Phosphates; D010716:Phosphatidylinositols; D017466:Receptors, Endothelin; D011983:Receptors, Purinergic; D019000:Phosphotyrosine; D000255:Adenosine Triphosphate; D005640:Follicle Stimulating Hormone; D000242:Cyclic AMP; D011493:Protein Kinase C; D010727:Phosphoric Diester Hydrolases; C021491:glycerophosphoinositol glycerophosphodiesterase; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D002118:Calcium