Intracellular Ca2+ responses and cell volume regulation upon cholinergic and purinergic stimulation in an immortalized salivary cell line.
The water channel aquaporin 5 (AQP5) seems to play a key role in salivary fluid secretion and appears to be critical in the cell volume regulation of acinar cells. Recently, the cation channel transient potential vanilloid receptor 4 (TRPV4) was shown to be functionally connected to AQP5 and also to cell volume regulation in salivary glands. We used the Simian virus 40 (SV40) immortalized cell line SMG C10 from the rat submandibular salivary gland to investigate the effect of ATP and the neurotransmitter analogue carbachol on Ca(2+) signalling and cell volume regulation, as well as the involvement of TRPV4 in the responses. We used fura-2-AM imaging, cell volume measurements, and western blotting. Both carbachol and ATP increased the concentration of intracellular Ca(2+), but no volume changes could be measured. Inhibition of TRPV4 with ruthenium red impaired both ATP- and carbachol-stimulated Ca(2+) signals. Peak Ca(2+) signalling during hyposmotic exposure was significantly decreased following inhibition of TRPV4, while the cells' ability to volume regulate appeared to be unaffected. These results show that in the SMG C10 cells, simulation of nervous stimulation did not induce cell swelling, although the cells had intact volume regulatory mechanisms. Furthermore, even though Ca(2+) signals were not needed for this volume regulation, TRPV4 seems to play a role during ATP and carbachol stimulation.
Aure
Marit H
MH
Department of Oral Biology, Faculty of Dentistry, University of Oslo, Blindern, Oslo, Norway.
Røed
Asbjørn
A
Galtung
Hilde Kanli
HK
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D051402:Aquaporin 5 / Q000187:drug effects; D001285:Atropine / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects*; D002217:Carbachol / Q000494:pharmacology; D018929:Cell Culture Techniques; D002461:Cell Line, Transformed; D048429:Cell Size / Q000187:drug effects; D018678:Cholinergic Agents / Q000494:pharmacology*; D018679:Cholinergic Agonists / Q000494:pharmacology; D005456:Fluorescent Dyes; D016257:Fura-2 / Q000031:analogs & derivatives; D018727:Muscarinic Antagonists / Q000494:pharmacology; D009997:Osmotic Pressure / Q000502:physiology; D058919:Purinergic P2 Receptor Antagonists; D011687:Purines / Q000494:pharmacology*; D011732:Pyridoxal Phosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D051381:Rats; D058469:Receptors, Purinergic P2X; D012430:Ruthenium Red / Q000494:pharmacology; D013363:Submandibular Gland / Q000166:cytology / Q000187:drug effects*; D013498:Suramin / Q000494:pharmacology; D050916:TRPV Cation Channels / Q000037:antagonists & inhibitors / Q000187:drug effects
C496040:Aqp5 protein, rat; D051402:Aquaporin 5; D018678:Cholinergic Agents; D018679:Cholinergic Agonists; D005456:Fluorescent Dyes; D018727:Muscarinic Antagonists; D058919:Purinergic P2 Receptor Antagonists; D011687:Purines; D058469:Receptors, Purinergic P2X; D050916:TRPV Cation Channels; C486577:Trpv4 protein, rat; C049925:fura-2-am; D012430:Ruthenium Red; C077792:pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid; D011732:Pyridoxal Phosphate; D013498:Suramin; D001285:Atropine; D000255:Adenosine Triphosphate; D002217:Carbachol; D016257:Fura-2