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We examined intracellular Ca2+ responses of the nasal gland acinar cells to clarify cellular responses and molecular events with regard to the regulatory mechanism of the nasal secretion. The acinar cells of the serous gland in the nasal septum of guinea pigs were obtained by meticulous and selective dissection with minimal contamination by epithelial lining cells after collagenase treatment. The dispersed acini were incubated in the oxygenated solution supplemented with fura 2 acetoxymethyl ester, and the intracellular Ca2+ concentration ([Ca2+]i) was measured by fluorescence ratio imaging microscopy. The application of acetylcholine (ACh) to the cells induced an initially rapid increased [Ca2+]i followed by a sustained plateau. The increase in [Ca2+]i induced by ACh was concentration dependent, ranging between 10(-8) and 10(-4) M. The [Ca2+]i response was completely inhibited by atropine, further indicating the involvement of muscarinic cholinergic receptors. Removal of external Ca2+ with addition of EGTA resulted in a transient increase without a sustained phase, and the transient increase was abolished by the intracellular Ca2+ antagonist 8-(diethylamino)-octyl-3,4,5-trimethoxybenzoate, indicating that this increase in [Ca2+]i was due to release from internal stores. The initial peak was not altered by changes in external pH, addition of adenosine 3',5'-cyclic monophosphate (cAMP), nor addition of phorbol 12-myristate 13-acetate (PMA) but was augmented by external K(+)-induced depolarization, suggesting that the transient increase was due to a changing in the binding affinity to inositol 1,4,5-trisphosphate. The sustained Ca2+ entry induced by ACh was inhibited by Ni2+, but not by nifedipine.(ABSTRACT TRUNCATED AT 250 WORDS)

Sho-seiryu-to, a traditional Japanese herbal medicine, has been used extensively in the treatment of allergic rhinitis. The effects of Sho-seiryu-to on electrical responses induced by acetylcholine in dissociated nasal gland acinar cells were investigated using patch-clamp and microfluorimetric imaging techniques. The application of Sho-seiryu-to inhibited both K+ and Cl- currents augmented by acetylcholine. The elevation of intracellular Ca2+ and Na+ concentrations induced by acetylcholine was also inhibited by Sho-seriyu-to. These findings suggest that Sho-seiryu-to attenuated the secretion of water and electrolytes from the nasal glands through an anti-cholinergic effect.

A procedure for isolating the acinar cells of the serous gland in the mammalian nasal septum has been developed. This technique is characterized by meticulous and selective isolation with minimal contamination by the surface epithelial cells and employs enzymatic treatment with collagenase. The isolated cells were confirmed to be serous gland acini as shown by negative staining with Alcian blue and a high electron density of the granules. The acini were more than 90% viable as judged by trypan blue exclusion. Ultrastructural integrity of the cells was well maintained following the isolation procedure. Application of acetylcholine to the isolated acini induced an inward current in a whole-cell patch clamp and increased intracellular Ca2+ concentration measured by fura-2. These acetylcholine responses were completely blocked by atropine. These physiological findings directly demonstrated that nasal gland acini possess muscarinic-activated receptors as previously suggested. These isolated cells hold promise for the in vitro study of secretory mechanisms in the mammalian nasal gland.

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