Calcium mobilization by activation of M(3)/M(5) muscarinic receptors in the human retinoblastoma.
Activation of muscarinic acetylcholine receptors (mAChR) is one of the most important signal transduction pathways in the human body. In this study, we investigated the role of mAChR activation in relation to its subtypes in human retinoblastoma cell-lines (WERI-Rb-1) using Ca(2+) measurement, real-time PCR, and Western Blot techniques. Acetylcholine (ACh) produced prominent [Ca(2+)](i) transients in a repeated manner in WERI-Rb-1 cells. The maximal amplitude of the [Ca(2+)](i) transient was almost completely suppressed by 97.3 +/- 0.8% after atropine (1 microM) pretreatment. Similar suppressions were noted after pretreatments with thapsigargin (1 microM), an ER Ca(2+)-ATPase (SERCA) inhibitor, whereas the ACh-induced [Ca(2+)](i) transient was not affected even in the absence of extracellular calcium. U-73122 (1 microM), a PLC inhibitor, and xestospongin C (2 microM), an IP(3)-receptor antagonist, elicited 11.5 +/- 2.9% and 17.8 +/- 1.9% suppressions, respectively. The 50% inhibitory concentration of (IC(50)) values for blockade of a 100 microM ACh response by pirenzepine and 4-DAMP were 315.8 and 9.1 nM, respectively. Moreover, both M(3) and M(5) mAChRs were prominent in quantitative real-time-PCR. Taken together, the M(3)/M(5) subtypes appear to be the major contributor, leading to intracellular calcium mobilization from the internal store via an IP(3)-dependent pathway in the undifferentiated retinoblastoma cells.
Kim
Dae-Ran
DR
Department of Basic Nursing Science and Institute for Nursing Science, Keimyung University College of Nursing, Daegu, Korea.
Rah
Sang Hoon
SH
Sohn
Joon Hyung
JH
Yeh
Byung-Il
BI
Ko
Chang Mann
CM
Park
Jeong Sook
JS
Kim
Min-Jeong
MJ
Lee
Joong Woo
JW
Kong
In Deok
ID
D000109:Acetylcholine / Q000008:administration & dosage / Q000494:pharmacology; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D045744:Cell Line, Tumor; D018678:Cholinergic Agents / Q000008:administration & dosage / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression; D006801:Humans; D043587:Receptor, Muscarinic M3 / Q000378:metabolism*; D043642:Receptor, Muscarinic M5 / Q000378:metabolism*; D012175:Retinoblastoma / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018678:Cholinergic Agents; D043587:Receptor, Muscarinic M3; D043642:Receptor, Muscarinic M5; D000109:Acetylcholine; D002118:Calcium