Regulation by isoproterenol of muscarinic acetylcholine receptor numbers and sensitivity in rat submandibular, but not lacrimal, glands

Chronic administration of -isoproterenol, a [beta]-adrenergic agonist, to male Sprague-Dawley rats increased submandibular gland weights by 3 to 4-fold. This increase resulted from a combination of hyperplasia and hypertrophy of secretory cells. Possible effects of this drug regimen on submandibular gland muscarinic acetylcholine receptors were examined by analysis of the binding of the cholinergic antagonist, -quinuclidinyl [3H]benzilate, to receptors in gland homogenates. Parallel investigations of receptors in exorbital lacrimal glands, an organ that is not grossly affected by chronic isoproterenol treatment, were also carried out. [3H]QNB bound to submandibular receptors with a Kd of 37.8+/-6.3 pM in control rats and 41.0+/-4.0 pM in isoproterenol-treated animals, a non-significant difference (P > 0.05). In contrast, the maximal binding level (Bmax) is isoproterenol-treated rats, 1.52+/-0.10 fmol/[mu]g DNA, was depressed by approx. 30% (PKd (61.3+/-5.3 vs. 53.2+/-4.0 pM) and Bmax (1.74+/-0.24 vs. 1.78+/-0.17 fmol/[mu]g DNA) were unchanged by isoproterenol treatment. The affinity of glandular muscarinic receptors for cholinergic agonists was also examined by competition experiments using carbachol. This agonist inhibited [3H]QNB binding to receptors in homogenates from both glands in a dose-dependent fashion. Inhibition constant (Ki) for this interaction were similar in control and isoproterenol-treated lacrimal glands; 53.6+/-5.4 [mu]M and 66.6+/-7.9 [mu]M, respectively (P>0.05). In submandibular glands, isoproterenol treatment elicited a highly significant (P ) shift in Ki from 17.3+/-1.4 [mu]M to 68.3+/-5.2 [mu]M. These results demonstrate that chronic administration of isoproterenol to rats results in a reduction in receptor numbers and a decrease in their sensitivity to cholinergic agonists in submandibular, but not lacrimal, glands.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/23949/1/0000196.pd

Fine structure and function of the neck organ in the brine shrimp Artemia salina nauplius

The morphology of the dorsal neck organ of the brine shrimp Artemia salina nauplius was investigated to determine its role(s) during larval development. Light and scanning electron microscopic studies show that the neck organ is present in the pre-nauplius larval forms as early as 12 hours after hydration of the desiccated cysts. It reaches its maximum development by the second or third instar but dedifferentiates in later stages as the phyllopodia mature. Areas of chloride ion accumulation in the E-2 and nauplius were localized by silver nitrate staining, The neck organ was the only portion of the larvae that stained to an appreciable degree, suggesting that it is the major site of sodium chloride secretion in the larval forms. Ultrastructural study of the neck organ in the early nauplius indicates that the organ serves the larva as a salt-secretory gland. The cells of the neck organ exhibit fine structural specializations commonly associated with epithelia involved in ion transport. The apical surfaces of the cells are extensively convoluted and the cell cytoplasm is divided into a web-like labyrinth which contains numerous mitochondria. Electron microscopic examination of the neck organ also reveals that it serves as a stable point of attachment for the dorsal antennal musculature, thus compensating for the non-rigid nature of the larval cuticle. It is suggested that the unusual architecture of the neck organ in the nauplius of Artemia may be an adaptation related to efficiency in both salt secretion and locomotion

Effects of cycloheximide and tunicamycin on cell surface expression of pancreatic muscarinic acetylcholine receptors

AbstractThe importance of glycosylation in cell surface expression of muscarinic receptors in cultured guinea pig pancreatic acini was investigated. Recovery of the muscarinic receptor population after carbachol-induced down regulation was blocked by cycloheximide but not by tunicamycin, although tunicamycin reduced [3H]mannose incorporation into acinar macromolecules by up to 90%. Tunicamycin treatment also failed to alter carbachol stimulation of amylase secretion from cultured acini. These results indicate that glycosylation of the glandular subtype of muscarinic receptor in the pancreatic acinar cell is not necessary for its insertion in the plasma membrane or for its functional activity

Secretagogue effects on intracellular calcium in pancreatic duct cells

Regulation of intracellular free calcium ([Ca 2+ ] i ) in single epithelial duct cells of isolated rat and guinea pig pancreatic interlobular ducts by secretin, carbachol and cholecystokinin was studied by microspectrofluorometry using the Ca 2+ -sensitive, fluorescent probe Fura-2. Rat and guinea pig duct cells exhibited mean resting [Ca 2+ ] i of 84 nM and 61 nM, respectively, which increased by 50%–100% in response to carbachol stimulation, thus demonstrating the presence of physiologically responsive cholinergic receptors in pancreatic ducts of both species. The carbachol-induced increase in [Ca 2+ ] i involved both mobilization of Ca 2+ from intracellular stores and stimulation of influx of extracellular Ca 2+ . In contrast, neither cholecystokinin nor secretin showed reproducible or sizeable increses in [Ca 2+ ] i . Both rat and guinea pig duct cells showed considerable resting Ca 2+ permeability. Lowering or raising the extracellular [Ca 2+ ] i led, respectively, to a decrease or increase in the resting [Ca 2+ ] i . Application of Mn 2+ resulted in a quenching of the fluorescence signal indicating its entry into the cell. The resting Ca 2+ and Mn 2+ permeability could be blocked by La 3+ suggesting that it is mediated by a Ca 2+ channel.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/47456/1/424_2004_Article_BF00370610.pd

Muscarinic stimulation of phospholipid turnover in dissociated avian salt gland cells

Addition of carbamylcholine to 32P-prelabeled dissociated avian salt gland cells resulted in increased turnover of phosphatidic acid, phosphatidylinositol, phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate, which could be prevented by the inclusion of atropine. Carbamylcholine had no discernable effect on protein phosphorylation, measured either in the total preparation or in subcellular fractions. It is concluded that for the avian salt gland, no obligatory link is indicated between protein phosphorylation and either phospholipid turnover or salt secretion.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/25220/1/0000660.pd

Freeze-fracture and morphometric analysis of occluding junctions in rectal glands of elasmobranch fish

The structure of occluding junctions in secretory and ductal epithelium of salt-secreting rectal glands from two species of elasmobranch fish, the spiny dogfish Squalus acanthias and the stingray Dasyatis sabina , was examined by thin-section and freeze-fracture electron microscopy. In both species, occluding junctions between secretory cells are shallow in their apical to basal extent and are characterized by closely juxtaposed parallel strands. Average strand number in the dogfish was 3.5±0.2 with a mean depth of 56±5 nm; in the stingray a mean of 2.0±0.2 strands encompassed an average depth of 18±3 nm. In contrast, the linear extent of these junctions was remarkably large due to the intermeshing of the narrow apices of the secretory cells to form the tubular lumen. Morphometric analysis gave values of 66.8±2.5 and 74.9±4.6 m/cm 2 for the length of junction per unit of luminal surface area in the dogfish and stingray, respectively. This junctional morphology is similar to that generally described for “leaky” epithelia. In comparison, the stratified ductal epithelium which carries the NaCl-rich secretion to the intestine is characterized by extensive occluding junctions which extend 0.6–0.8 μm in depth and consist of a mean of 12 strands arranged in an anastomosing network, an architectural pattern typical of “tight” epithelia. The length density of these junctions in the dogfish rectal gland was 7.6±0.1 m/cm 2 .Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/48030/1/232_2005_Article_BF01870973.pd