Naoto Asada

Salivary Secretion of Highly Concentrated Chromogranin a in Response to Noradrenaline and Acetylcholine in Isolated and Perfused Rat Submandibular Glands

Chromogranin A (CgA) is a member of a family of highly acidic proteins, chromogranins, which are co‐stored in the adrenergic neurons and paraneurons and co‐released with adrenaline and noradrenaline (NAd) in response to adequate stimulation. The present study provides novel evidence that CgA‐like immunoreactivity (IR) is stored in the exocrine cells in the granular convoluted tubule, and is secreted into saliva by stimulation with NAd and acetylcholine (ACh) in the isolated and perfused rat submandibular gland. NAd at 1 μM produced maximum secretion of CgA‐like IR (<< 0.9 mM) and a marked increase in salivary flow. Further increases in NAd concentration (10 or 100 μM) yielded concentration‐dependent decreases in both responses. ACh at 1 μM produced maximum salivary flow and a slight elevation of CgA‐like IR secretion (6 μM); 100 μM ACh decreased the salivary flow but increased the CgA‐like IR secretion (0.6 mM). Electron microscopic examination showed vigorous compound exocytosis of secretory granules in the cells of the granular convoluted tubule when the submandibular gland was stimulated with 1 μM NAd. These results provide an experimental basis for the view that the salivary CgA‐like IR secretion may be a sensitive and quantitative index of the activity of the sympathetic nervous system innervating the gland.

Salivary secretion of chromogranin A. Control by autonomic nervous system.

We have provided novel evidence that CGA-LI is stored in the cells of granular convoluted tubule, and is secreted into saliva by stimulation with NAd in the isolated and perfused rat submandibular gland. The cells are also known to secrete various growth factors, which are released into blood stream. The cells may release CGA-LI into blood stream (endocrine secretion) in addition to salivary release (exocrine secretion). The endocrine- exocrine secretion of the cells resembles the secretion of renin in the cells of juxtaglomerular apparatus and coagulating gland.

VIP‐ and PACAP‐Induced Salivary Chromogranin A Secretion in the Isolated Perfused Submandibular Gland of Rats

Abstract: In the study reported in this paper, sensitive ELISA for rat CgA was developed using synthetic rat CgA(359–389) as antigen, Nα‐biotinylated glycylglycyl rat CgA(359–389), and antirat CgA(359–389) serum for the measurement of CgA‐LI in rat saliva. CgA‐LI in rat submandibular tissues and saliva was characterized by both immunohistochemical and immunochemical methods. Using isolated perfused rat submandibular gland. VIP at 0.1–1.0 nM in the presence of 0.1 μM ACh was found to cause CgA‐LI secretion, whereas neither PACAP‐27 nor PACAP‐38 showed any effect on CgA secretion.

Acidification of Extracellular Environment Inhibits CCK-8-Induced Ca2+ Entry into Pancreatic Acinar Cells of Rat Evaluated by a Mn2+-Quenching Method

The fluorescence of fura 2, a Ca(2+)-sensitive dye, is quenched in the presence of other divalent cations such as Mn2+. This characteristic of the dye provides a useful measure for investigating the role of Ca2+ in cellular functions. The present experiments were thus designed to examine the effects of extracellular pH (pHo) on cholecystokinin octapeptide (CCK-8)-induced Ca2+ entry into isolated perifused pancreatic acini of the rat using a Mn(2+)-quenching method. At a standard pHo (7.4), addition of Mn2+ (1 mM) during continuous stimulation with 100 pM CCK-8 quenched fura 2 fluorescence excited by both excitation wavelengths of 340 and 380 nm. Lowering pHo to 6.0 weakened the quenching, whereas elevating pHo to 8.0 during the stimulation accelerated the quenching compared with that at standard pHo. Accordingly, when acinar cells were stimulated continuously with 100 pM CCK-8 at pHo 6.0, the second sustained increase in cytosolic Ca2+ concentration ([Ca2+]i) was decreased compared with that at pHo 7.4, but the [Ca2+]i increase reappeared when the pHo was neutralized from 6.0 to 7.4 even after cessation of the stimulation. The second sustained increase of [Ca2+]i was achieved at pHo 8.0, but it declined rapidly when CCK-8 stimulation was terminated and pHo was simultaneously restored to 7.4. Amylase release induced by continuous stimulation with 100 pM CCK-8 showed a biphasic time course. A second sustained phase of amylase release was significantly reduced at pHo 6.0, but amylase release increased again when pHo was restored to 7.4. These results indicate that extracellular pH modifies the extent of secretagogue-induced Ca2+ entry into the pancreatic acinar cell, and the inhibitory effects of lowered pHo on the secretory response are largely due to a limited increase in [Ca2+]i.