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The G protein-coupled cholecystokinin 1 receptor (CCK1R) is activated permanently by type II photodynamic action (i.e., by singlet oxygen) in the freshly isolated rat pancreatic acini, in contrast to reversible activation by CCK. But how CCK1R is photodynamically activated is not known. Therefore, in the present work, we subjected membrane proteins extracted from isolated rat pancreatic acini to photodynamic action with photosensitiser sulphonated aluminium phthalocyanine (SALPC), and used reducing gel electrophoresis and Western blot to detect possible changes in CCK1R oligomerization status. Photodynamic action (SALPC 1 µM, light 36.7 mW cm - 2  × 10 min) was found to convert dimeric CCK1R nearly quantitatively to monomers. Such conversion was dependent on both irradiance (8.51-36.7 mW cm - 2 ) and irradiation time (1-20 min). Minimum effective irradiance was found to be 11.1 mW cm - 2 (× 10 min, with SALPC 1 µM), and brief photodynamic action (SALPC 1 µM, 36.7 mW cm - 2  × 1 min) was effective. Whilst CCK stimulation of purified membrane proteins alone had no effect on CCK1R dimer/monomer balance, sub-threshold photodynamic action (SALPC 100 nM, 36.7 mW cm - 2  × 10 min) plus CCK revealed a bell-shaped CCK dose response curve for CCK1R monomerization, which was remarkably similar to the dose response curve for CCK-stimulated amylase secretion in isolated rat pancreatic acini. These two lines of evidence together suggest that during photodynamic CCK1R activation, CCK1R is permanently monomerized, thus providing a unique approach for permanent G protein-coupled receptor (GPCR) activation which has not been achieved before.

Solar UVA radiation (320-400 nm) is known to have immunomodulatory effects, but the detailed mechanisms involved are not fully elucidated. UVA irradiation has been shown to induce calcium oscillations in rat peritoneal mast cells due to NAD(P)H oxidase (NOX) activation, but the specific NOX isoforms have not been identified. In the present work effects of UVA irradiation were investigated in isolated rat peritoneal mast cells, in cultured rat mast cell line RBL-2H3, and in mouse bone marrow-derived mast cells (BMMC). It was found that UVA irradiation by alternate 340/380 nm (3.2-5.6 μW cm(-2)) or by LED (380 nm, 80 μW cm(-2)) induced calcium oscillations in isolated rat peritoneal mast cells, in RBL-2H3, and in BMMC. Such UVA-induced calcium oscillations resembled closely those induced by surface IgE receptor (FcεRI) activation. It was found that RBL-2H3 expressed high levels of gp91(phox) (NOX2), p22(phox), p67(phox), p47(phox), p40(phox), Rac1, Rac2, moderate levels of DUOX2, but did not express NOX1, NOX3, NOX4, or DUOX1. The specific cellular localizations of gp91(phox) (NOX2), p22(phox), p47(phox), p67(phox), p40(phox) and Rac1/2 were confirmed by immunocytochemistry. UVA-induced reactive oxygen species (ROS) production in RBL-2H3 was completely suppressed by the NOX inhibitor diphenyleneiodonium chloride (DPI) or by the antioxidant N-acetyl-l-cysteine (NAC). siRNA suppression of gp91(phox) (NOX2), p22(phox) and p47(phox) expression inhibited markedly UVA-induced calcium oscillations, ROS and IL-6/LTC4 production in RBL-2H3. Taken together these data indicate that NOX2 plays an essential role in UVA irradiation-induced calcium oscillations, ROS and mediator production in mast cells.

Toosendanin, a triterpenoid from Melia toosendan Sieb et Zucc, has been found before to be an effective anti-botulism agent, with a bi-phasic effect at both motor nerve endings and central synapse: an initial facilitation followed by prolonged depression. Initial facilitation may be due to activation of voltage-dependent calcium channels plus inhibition of potassium channels, but the depression is not fully understood. Toosendanin has no effect on intracellular calcium or secretion in the non-excitable pancreatic acinar cells, ruling out general toosendanin inhibition of exocytosis. In this study, toosendanin effects on sensory neurons isolated from rat nodose ganglia were investigated. It was found that toosendanin stimulated increases in cytosolic calcium and neuronal exocytosis dose dependently. Experiments with membrane potential indicator bis-(1,3-dibutylbarbituric acid)trimethine oxonol found that toosendanin hyperpolarized capsaicin-insensitive but depolarized capsaicin-sensitive neurons; high potassium-induced calcium increase was much smaller in hyperpolarizing neurons than in depolarizing neurons, whereas no difference was found for potassium-induced depolarization in these two types of neurons. In neurons showing spontaneous calcium oscillations, toosendanin increased the oscillatory amplitude but not frequency. Toosendanin-induced calcium increase was decreased in calcium-free buffer, by nifedipine, and by transient receptor potential vanilloid 1 (TRPV1) antagonist capsazepine. Simultaneous measurements of cytosolic and endoplasmic reticulum (ER) calcium showed an increase in cytosolic but a decrease in ER calcium, indicating that toosendanin triggered ER calcium release. These data together indicate that toosendanin modulates sensory neurons, but had opposite effects on membrane potential depending on the presence or absence of capsaicin receptor/TRPV 1 channel.

This paper investigated the role of acetylcholine (ACh) in physiological regulation of amylase secretion in avian exocrine pancreas. In the isolated duck pancreatic acini, ACh dose dependently stimulated amylase secretion, with a maximal effective concentration at 10 muM. The cAMP-mobilizing compounds forskolin, vasoactive intestinal peptide (VIP)/pituitary adenylate cyclase activating peptide (PACAP) receptor (VPAC) agonists PACAP-38 and PACAP-27 had no effect on the dose-response curve. ACh dose dependently induced increases in cytosolic Ca(2+) concentration ([Ca(2+)]( c )), with increasing concentrations transforming oscillations into plateau increases. Forskolin (10 muM), PACAP-38 (1 nM), PACAP-27 (1 nM), or VIP (10 nM) alone did not stimulate [Ca(2+)]( c ) increase; neither did they modulate ACh-induced oscillations, nor made ACh low concentration effective. These data indicate that ACh-stimulated zymogen secretion in duck pancreatic acinar cells is not subject to modulation from the cAMP signaling pathway; whereas it has been widely reported in the rodents that ACh-stimulated exocrine pancreatic secretion is significantly enhanced by cAMP-mobilizing agents. This makes the duck exocrine pancreas unique in that cholinergic stimulus-secretion coupling is not subject to cAMP regulation.

The adrenergic agonist phenylephrine (PE) induced typical [Ca2+]i oscillations in freshly isolated rat hepatocytes, and the photodynamic effect of Victoria Blue BO on induced calcium oscillations was investigated. PE induced calcium oscillations disappeared after Victoria Blue BO photodynamic action (in 20 out of 26 experiments), or were present with a decreased amplitude (in 1 out of 26 experiments) or frequency (in 5 out of 26 experiments). After VBBO photodynamic action, the basal calcium level remained at pre-stimulatory level. In comparison, after treatment with the mitochondrial uncoupler CCCP. calcium oscillations disappeared in all experiments, with a significant increase in basal [Ca2+]i. After washing out of CCCP, basal [Ca2+]i returned to pre-stimulation level and calcium oscillations recovered. The photodynamically-induced blockade of calcium oscillations was irreversible, and this effect was present independent of the time interval between VBBO incubation and light illumination. Calcium oscillation blockade was not seen with VBBO in the dark at the concentration used, nor was it seen with light illumination alone. Taken together. it is concluded that VBBO photodynamic action could irreversibly block calcium oscillations in rat hepatocytes, possibly due to blockade of mitochondrion calcium release.