Calcium by a Phospholipase C-Dependent Mechanism

Calcium signaling in fura-2 acetoxymethyl ester-loaded enteric glia was investigated in response to neuroligands; responses to AlP were studied in detail. Carbachol (1 mM), glutamate (100 jiM), norepinephrine (10 jiM), and substance P (1 pM) did not increase the intracellular calcium concentration ([Ca24 ],) in cultured enteric glia. An increasing percentage of glia responded to serotonin (4%; 100 pM), bradykinin (11%; 10 1eM), and histamine (31%; 100 pM), whereas 100% of glia responded to ATP (100 pM). ATP-evoked calcium signaling was concentration dependent in terms of the percentage of glia responding and the peak [Ca 2fl, achieved; responses w re pertussis toxin ins nsitive. Based on responsiveness of enteric glia to purinergic agonists and peak [Ca2~]evoked, ATP UTP > ADP > j3,y-methyleneadenosine 5 ‘-triphosphate ©‘ 2-methylthioadenosine 5 ‘-triphosphate = ev,fl-methyleneadenosine 5 ‘-triphosphate = AMP = adenosine, suggesting a glial P 2~receptor. Depletion of D-myo-inositol 1 ,4,5-trisphosphate-sensitive calcium stores by thapsigargin (10 pM) abolished glial responses to ATP. Similarly, calcium responses were decreased 92% by U-73122 (10 pM), an inhibitor of phospholipase C, and 93% by the phorbol ester phorbol 12-myristate 13-acetate (100 nM), an activator of protein kinase C. Thus, cultured enteric glia can respond to neurotransmitters with increases in [Ca 2~}. Our data sugg st that glial responses to ATP are mediated by a Psu receptor coupled to activation of phospholipase C and release of intracellular calcium stores.