Latency in the inositol lipid transduction pathway: the role of cellular events in responses to thyrotropin-releasing hormone in Xenopus oocytes.
To dissect the cellular events responsible for the prolonged latency of the response to thyrotropin-releasing hormone (TRH) in Xenopus oocytes we interfered with different steps of the signal transduction pathway. Preincubation of oocytes with cis-vaccenic acid (a membrane-fluidizing agent) shortened the latency, suggesting a contribution of membranal processes. TRH-induced depletion of cellular calcium stores prolonged latency (up to threefold), which returned to control levels upon repletion of the stores. Injection of D-2,3-diphosphoglycerate (PGA), which inhibits inositol (1,4,5)-trisphosphate (InsP3) dephosphorylation, alone evoked a small, prolonged depolarizing current and significantly shortened the latency of the response to TRH. Injection of guanosine 5'-O-(2-thiodiphosphate) (GDP beta S), which inactivates guanine nucleotide-binding regulatory proteins, decreased the amplitude of the response and increased latency. Injection of guanosine 5-O-(3-thiotriphosphate) (GTP gamma S) immediately before the challenge with TRH did not shorten the latency of the response. Decreasing the effective receptor density with chlordiazepoxide, an antagonist of the TRH receptor, resulted in an extension of latency, whereas the expression of a large number of TRH receptors by injection of RNA transcribed from cloned receptor DNA (10-100 ng/oocyte) shortened the latency to below 2 s. Our results suggest that the latency of the response to TRH reflects the activation of a late step in the signal transduction sequence, most likely the release of calcium by InsP3. We propose that this process is kinetically controlled by an early rate-limiting event, involving the activation of a guanine nucleotide-binding protein by the TRH receptor.
Lipinsky
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D
Department of Physiology and Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Israel.
Gershengorn
M C
MC
Oron
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Y
D019794:2,3-Diphosphoglycerate; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D004163:Diphosphoglyceric Acids / Q000494:pharmacology; D005260:Female; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D009829:Oleic Acids / Q000494:pharmacology; D009865:Oocytes / Q000187:drug effects / Q000502:physiology*; D018025:Receptors, Thyrotropin-Releasing Hormone / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D013873:Thionucleotides / Q000494:pharmacology; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology*; D013997:Time Factors; D014981:Xenopus
D004163:Diphosphoglyceric Acids; D009829:Oleic Acids; D018025:Receptors, Thyrotropin-Releasing Hormone; D013873:Thionucleotides; D019794:2,3-Diphosphoglycerate; D006153:Guanosine Diphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); C065593:cis-vaccenic acid; D013973:Thyrotropin-Releasing Hormone; C023427:guanosine 5'-O-(2-thiodiphosphate); D015544:Inositol 1,4,5-Trisphosphate; D002118:Calcium