Ca2+ signaling in prothoracicotropic hormone-stimulated prothoracic gland cells of Manduca sexta: evidence for mobilization and entry mechanisms.
Prothoracicotropic hormone (PTTH) stimulates ecdysteroidogenesis in lepidopteran prothoracic glands (PGs), thus indirectly controlling molting and metamorphosis. PTTH triggers a signal transduction cascade in PGs that involves an early influx of Ca2+. Although the importance of Ca2+ has been long known, the mechanism(s) of PTTH-stimulated changes in cytoplasmic Ca2+ [Ca2+]i are not yet well understood. PGs from the fifth instar of Manduca sexta were exposed to PTTH in vitro. The resultant changes in [Ca2+]i were measured using ratiometric analysis of a fura-2 fluorescence signal in the presence and absence of inhibitors of specific cellular signaling mechanisms. The phospholipase C (PLC) inhibitor U-73122 nearly abolished the PTTH-stimulated increase in [Ca2+]i, as well as PTTH-stimulated ecdysteroidogenesis and extracellular-signal regulated kinase phosphorylation, thus establishing a role for PLC and implicating inositol trisphosphate (IP3) in PTTH signal transduction. Two antagonists of the IP3 receptor, 2-APB and TMB-8, likewise blocked the [Ca2+]i response by a mean of 92%. We describe for the first time the presence of Ca2+ oscillations in PTTH-stimulated cells in Ca2+-free medium. External Ca2+ entered PG cells via at least two routes: store-operated (capacitative) Ca2+ entry channels and L-type voltage-gated Ca2+ channels. We propose that PTTH initiates a transductory cascade typical of many G-protein coupled receptors, involving both Ca2+ mobilization and entry pathways.
Fellner
Susan K
SK
Department of Cell and Molecular Physiology (CB# 7545), University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Rybczynski
Robert
R
Gilbert
Lawrence I
LI
D000818:Animals; D020013:Calcium Signaling / Q000502:physiology*; D004441:Ecdysterone / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006023:Glycoproteins / Q000494:pharmacology; D007301:Insect Hormones / Q000494:pharmacology*; D007814:Larva / Q000187:drug effects / Q000502:physiology; D018402:Manduca / Q000187:drug effects / Q000254:growth & development / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013909:Thorax / Q000502:physiology
D006023:Glycoproteins; D007301:Insect Hormones; D004441:Ecdysterone; C009089:prothoracicotropic hormone; D048049:Extracellular Signal-Regulated MAP Kinases