Co-contribution of IP3R and Ca2+ influx pathways to pacemaker Ca2+ activity in stomach ICC.
Intracellular Ca2+ oscillations in interstitial cells of Cajal (ICCs) are thought to be the primary pacemaker activity in the gut. In the present study, the authors prepared small tissues of 100-to 300-microm diameter (cell cluster preparation) from the stomach smooth muscle (including the myenteric plexus) of mice by enzymatic and mechanical treatments. After 2 to 4 days of culture, the intracellular Ca2+ concentration ([Ca2+]i) was measured. In the presence of nifedipine, a dihydropyridine Ca2+ channel antagonist, spontaneous [Ca2+]i oscillations were observed within limited regions showing positive c-Kitimmunoreactivity, a maker for ICCs. In the majority of cell cluster preparations with multiple regions of [Ca2+]i oscillations, [Ca2+]i oscillated synchronously in the same phase. A small number of cell clusters (8 of 53) showed multiple regions of [Ca2+]i oscillations synchronized but with a considerable phase shift. Neither tetrodotoxin (250 nM) nor atropine (10 microM) significantly affected [Ca2+]i oscillations in the presence of nifedipine. Low concentrations (40 microM) of Ni2+ had little effect on the spontaneous [Ca2+]i oscillation, but SK&F96365 (40 microM) and Cd2+ (120 microM) terminated it. Applications of either 2-aminoethoxydiphenyl borate (10 microM) or xestosponginC(10 microM) completely and rather rapidly (approximately 2 min) abolished the spontaneous [Ca2+]i oscillations. The results suggest that pacemaker [Ca2+]i oscillations in ICCs are produced by close interaction of intracellular Ca2+ release channels, especially inositol 1,4,5-trisphosphate receptor (InsP3R) and Ca2+ influx pathways, presumably corresponding to store-operated type channels. Reverse transcription polymerase chain reaction examinations revealed expression of TRPC2, 4, and 6, as well as InsP3R1 and 2 in ICCs.
Liu
Hong-Nian
HN
Department of Cell Physiology, Nagoya University Graduate School of Medicine, Nagoya, Japan.
Ohya
Susumu
S
Furuzono
Shinji
S
Wang
Jing
J
Imaizumi
Yuji
Y
Nakayama
Shinsuke
S
D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism* / Q000502:physiology; D015220:Calcium Channels / Q000378:metabolism* / Q000502:physiology; D017931:DNA Primers; D005753:Gastric Mucosa / Q000378:metabolism; D066298:In Vitro Techniques; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008562:Membrane Glycoproteins / Q000378:metabolism* / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C; D009130:Muscle, Smooth / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D012333:RNA, Messenger / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism* / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013270:Stomach / Q000187:drug effects / Q000502:physiology*; D013779:Tetrodotoxin / Q000494:pharmacology
D015220:Calcium Channels; D017931:DNA Primers; D053496:Inositol 1,4,5-Trisphosphate Receptors; C025389:Itpr1 protein, mouse; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D013779:Tetrodotoxin; D002118:Calcium