Intracellular calcium concentration in the inositol trisphosphate receptor type 1 knockout mouse.
Recently, mice with a disrupted inositol trisphosphate (IP3) receptor type 1 allele were produced by gene targeting. To examine the role of IP3 receptor type I in the regulation of intracellular calcium concentration ([Ca2+]i) of glomerular cells, [Ca2+]i was measured with fura 2-acetoxymethyl-ester in the superfused glomeruli from homozygous and wild-type mice. [Ca2+]i was determined in calcium-free medium before and after the addition of 10(-7) M endothelin-1 (ET-1) and 10(-6) M angiotensin II (AngII). The expression of mRNA of IP3 receptor isoforms and hormone receptors in the glomeruli from these animals also was measured by quantitative reverse transcription-PCR with specific primers for IP3 receptor isoforms (types 1, 2, and 3), AngII receptor type 1, and ET receptors (types A and B). In homozygous mutants, the shorter mRNA of IP3 receptor type 1, which lacks the first exon, is transcribed. Basal [Ca2+]i and the responses to ET-1 and AngII in homozygous mutants (ET-1, 55 +/- 7 nM to 73 +/- 7 nM; AngII, 66 +/- 6 to 91 +/- 8 nM) were significantly lower than those in the wild-type mice (ET-1, 93 +/- 13 nM to 162 +/- 13 nM; AngII, 87 +/- 7 to 147 +/- 9 nM; P < 0.05 for both hormones) without significant changes in mRNA expression of hormone receptors. The results with quantitative reverse transcription-PCR also revealed that mRNA expression of the IP3 receptor gene family was not significantly different between the two groups. The present study clearly shows that IP3 receptor type 1 plays a major role in the regulation of [Ca2+]i in the glomeruli and that lack of an isoform of IP3 receptor in the glomeruli does not induce expression of the other isoforms of the IP3 receptor.
Hayashi
M
M
Department of Internal Medicine, School of Medicine, Keio University, Tokyo, Japan. matuhiko@mc.med.keio.ac.jp
Monkawa
T
T
Yoshida
T
T
Sasamura
H
H
Matsumoto
M
M
Inoue
T
T
Mikoshiba
K
K
Saruta
T
T
D000199:Actins / Q000235:genetics / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D016232:Endothelins / Q000378:metabolism; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism*; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007678:Kidney Glomerulus / Q000166:cytology / Q000378:metabolism*; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008817:Mice, Mutant Strains; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000032:analysis; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D000199:Actins; D015220:Calcium Channels; D016232:Endothelins; D053496:Inositol 1,4,5-Trisphosphate Receptors; D012333:RNA, Messenger; D018160:Receptors, Cytoplasmic and Nuclear; D015544:Inositol 1,4,5-Trisphosphate; D002118:Calcium