title: Plasma and intracellular membrane inositol 1,4,5-trisphosphate receptors mediate the Ca(2+) increase associated with the ATP-induced increase in ciliary beat frequency.
abstract: An increase in intracellular free Ca(2+) concentration ([Ca(2+)](i)) has been shown to be involved in the increase in ciliary beat frequency (CBF) in response to ATP; however, the signaling pathways associated with inositol 1,4,5-trisphosphate (IP(3)) receptor-dependent Ca(2+) mobilization remain unresolved. Using radioimmunoassay techniques, we have demonstrated the appearance of two IP(3) peaks occurring 10 and 60 s after ATP addition, which was strongly correlated with a release of intracellular Ca(2+) from internal stores and an influx of extracellular Ca(2+), respectively. In addition, ATP-dependent Ca(2+) mobilization required protein kinase C (PKC) and Ca(2+)/calmodulin-dependent protein kinase II activation. We found an increase in PKC activity in response to ATP, with a peak at 60 s after ATP addition. Xestospongin C, an IP(3) receptor blocker, significantly diminished both the ATP-induced increase in CBF and the initial transient [Ca(2+)](i) component. ATP addition in the presence of xestospongin C or thapsigargin revealed that the Ca(2+) influx is also dependent on IP(3) receptor activation. Immunofluorescence and confocal microscopic studies showed the presence of IP(3) receptor types 1 and 3 in cultured ciliated cells. Immunogold electron microscopy localized IP(3) receptor type 3 to the nucleus, the endoplasmic reticulum, and, interestingly, the plasma membrane. In contrast, IP(3) receptor type 1 was found exclusively in the nucleus and the endoplasmic reticulum. Our study demonstrates for the first time the presence of IP(3) receptor type 3 in the plasma membrane in ciliated cells and leads us to postulate that the IP(3) receptor can directly trigger Ca(2+) influx in response to ATP.
authors.affiliation: Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, Alameda 340, Casilla 114-D, Santiago, Chile.
authors.forename: Nelson P
authors.initials: NP
authors.lastname: Barrera
authors.forename: Bernardo
authors.initials: B
authors.lastname: Morales
authors.forename: Manuel
authors.initials: M
authors.lastname: Villalón
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D002923:Cilia / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure; D006224:Cricetinae; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D005455:Fluorescent Antibody Technique; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007425:Intracellular Membranes / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D010057:Oviducts / Q000166:cytology / Q000502:physiology; D011493:Protein Kinase C / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000187:drug effects / Q000502:physiology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology
chemical_list: D015220:Calcium Channels; D004791:Enzyme Inhibitors; D053496:Inositol 1,4,5-Trisphosphate Receptors; D018160:Receptors, Cytoplasmic and Nuclear; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium