Gramicidin-perforated patch recording revealed the oscillatory nature of secretory Cl- movements in salivary acinar cells

Elevations of cytoplasmic free calcium concentrations ([Ca(2+)](i)) evoked by cholinergic agonists stimulate isotonic fluid secretion in salivary acinar cells. This process is driven by the apical exit of Cl(-) through Ca(2+)-activated Cl(-) channels, while Cl(-) enters the cytoplasm against its electrochemical gradient via a loop diuretic-sensitive Na(+)-K(+)-2Cl(-) cotransporter (NKCC) and/or parallel operations of Cl(-)-HCO(3)(-) and Na(+)-H(+) exchangers, located in the basolateral membrane. To characterize the contributions of those activities to net Cl(-) secretion, we analyzed carbachol (CCh)-activated Cl(-) currents in submandibular acinar cells using the "gramicidin-perforated patch recording configuration." Since the linear polypeptide antibiotic gramicidin creates monovalent cation-selective pores, CCh-activated Cl(-) currents in the gramicidin-perforated patch recording were carried by Cl(-) efflux via Cl(-) channels, dependent upon Cl(-) entry through Cl(-) transporters expressed in the acinar cells. CCh-evoked oscillatory Cl(-) currents were associated with oscillations of membrane potential. Bumetanide, a loop diuretic, decreased the CCh-activated Cl(-) currents and hyperpolarized the membrane potential. In contrast, neither methazolamide, a carbonic anhydrase inhibitor, nor elimination of external HCO(3)(-) had significant effects, suggesting that the cotransporter rather than parallel operations of Cl(-)-HCO(3)(-) and Na(+)-H(+) exchangers is the primary Cl(-) uptake pathway. Pharmacological manipulation of the activities of the Ca(2+)-activated Cl(-) channel and the NKCC revealed that the NKCC plays a substantial role in determining the amplitude of oscillatory Cl(-) currents, while adjusting to the rate imposed by the Ca(2+)-activated Cl(-) channel, in the gramicidin-perforated patch configuration. By concerting with and being controlled by the cation steps, the oscillatory form of secretory Cl(-) movements may effectively provide a driving force for fluid secretion in intact acinar cells.     

Pancreatic exocrine damage induced by subcutaneous injection of a low dosage of zinc

The aim of this study was to observe whether a low dosage of zinc induced mouse pancreatic injury. Dosages of zinc from 0.1 to 50 mg/kg were injected subcutaneously in mice, and plasma and pancreatic clinical parameters were observed 3–24 h after the injection. Plasma α-amylase activity increased 10 and 24 h after the injection of 25 or 50 mg/kg of zinc, whereas pancreatic α-amylase activity decreased 3 h after more than 5 mg/kg of zinc was injected. The activity recovered after 24 h except in the group injected with 50 mg/kg of zinc. The plasma glucose level did not change when less than 25 mg/kg of zinc was injected. The pancreatic zinc contents increased 3 h after more than 1 mg/kg of zinc was injected. The pancreatic metallothionein (MT) contents increased 6 h after the injection of 1 mg/kg of zinc. In addition, when more than 5 mg/kg of zinc was injected, the MT content increased at 3 h. In histochemical observations, cell damages such as fibrosis and necrosis were observed in pancreatic exocrine cells, but not in cells of Langerhans islets. From the present study, a single injection of a low dosage of zinc induces injury in pancreatic exocrine cells, but not endocrine cells.     

Calcium wave propagation in pancreatic acinar cells: functional interaction of inositol 1,4,5-trisphosphate receptors, ryanodine receptors, and mitochondria

In pancreatic acinar cells, inositol 1,4,5-trisphosphate (InsP(3))-dependent cytosolic calcium ([Ca(2+)](i)) increases resulting from agonist stimulation are initiated in an apical "trigger zone," where the vast majority of InsP(3) receptors (InsP(3)R) are localized. At threshold stimulation, [Ca(2+)](i) signals are confined to this region, whereas at concentrations of agonists that optimally evoke secretion, a global Ca(2+) wave results. Simple diffusion of Ca(2+) from the trigger zone is unlikely to account for a global [Ca(2+)](i) elevation. Furthermore, mitochondrial import has been reported to limit Ca(2+) diffusion from the trigger zone. As such, there is no consensus as to how local [Ca(2+)](i) signals become global responses. This study therefore investigated the mechanism responsible for these events. Agonist-evoked [Ca(2+)](i) oscillations were converted to sustained [Ca(2+)](i) increases after inhibition of mitochondrial Ca(2+) import. These [Ca(2+)](i) increases were dependent on Ca(2+) release from the endoplasmic reticulum and were blocked by 100 microM ryanodine. Similarly, "uncaging" of physiological [Ca(2+)](i) levels in whole-cell patch-clamped cells resulted in rapid activation of a Ca(2+)-activated current, the recovery of which was prolonged by inhibition of mitochondrial import. This effect was also abolished by ryanodine receptor (RyR) blockade. Photolysis of d-myo InsP(3) P(4(5))-1-(2-nitrophenyl)-ethyl ester (caged InsP(3)) produced either apically localized or global [Ca(2+)](i) increases in a dose-dependent manner, as visualized by digital imaging. Mitochondrial inhibition permitted apically localized increases to propagate throughout the cell as a wave, but this propagation was inhibited by ryanodine and was not seen for minimal control responses resembling [Ca(2+)](i) puffs. Global [Ca(2+)](i) rises initiated by InsP(3) were also reduced by ryanodine, limiting the increase to a region slightly larger than the trigger zone. These data suggest that, while Ca(2+) release is initially triggered through InsP(3)R, release by RyRs is the dominant mechanism for propagating global waves. In addition, mitochondrial Ca(2+) import controls the spread of Ca(2+) throughout acinar cells by modulating RyR activation.     

The basicoxal gland, a new exocrine structure in poneromorph ants (Hymenoptera, Formicidae)

The tegumental epithelium of the outer dorsolateral region in the proximal part of the coxae in the mid‐ and hindlegs of both workers and queens of the ants Odontomachus rixosus and O. simillimus is differentiated into a conspicuous and hitherto unknown exocrine gland. The glandular cells display a clear microvillar differentiation of their apical cell membrane, and are lined with the tegumental cuticle, which in this part contains crack‐like channels perpendicular to its surface, that carry the glandular secretions to the outside. Apical microvilli support the transport of substances, and contain an extension of tubular smooth endoplasmic reticulum in their centre. The function of the gland may be that of providing lubricant substances to the articulation region of the generally heavily sclerotized ponerine ant species. The gland is also found in several other ponerine and amblyoponine species, but not in the ectatommine species studied. The foreleg coxae lack a basicoxal gland in all species examined, which may be explained by the more limited articulation between the thorax and the coxae in the forelegs compared to the mid‐ and hindlegs.     

Histology of mental and caudal courtship glands in three genera of plethodontid salamanders (Amphibia: Plethodontidae)

Salamanders in the family Plethodontidae exhibit a unique tail‐straddle walk during courtship that can include the use of sexually dimorphic mental and caudal courtship glands. This study presents novel histological and fine structure data on mental glands and caudal courtship glands in Plethodon mississippi, Desmognathus conanti and Eurycea quadridigitata using both light microscopy and scanning electron microscopy. This study represents the first use of scanning electron microscopy to observe these glands. Both mental and caudal courtship glands were observed to vary seasonally in gland diameter and histology according to the breeding season of each species. Morphological variation was observed across the three species studied in both clustering and relative size of the glands compared to neighbouring mucous and granular glands. Hypertrophied mental glands are larger than mucous or granular glands in all species, but relationships among caudal courtship glands and other skin glands vary among species. In E. quadridigitata, active caudal courtship glands are larger than mucous and granular glands, but in D. conanti, caudal courtship glands are similar in size to granular glands and larger than mucous glands. In P. mississippi, caudal courtship glands are scattered among significantly larger granular glands and are similar in size to mucous glands.     

Effect of pancreatic polypeptide-antiserum on bombesin-stimulated pancreatic exocrine secretion in dogs

Bombesin is a potent stimulus of both pancreatic protein secretion and plasma pancreatic polypeptide (PP) release in dogs. Physiological plasma levels of PP have been shown to inhibit pancreatic exocrine secretion in dogs. We examined the question whether the concomitant release of PP exerts a suppressive action on the pancreatic exocrine response to bombesin in dogs by measuring pancreatic exocrine secretion with and without in vivo immunoneutralization of PP with a high affinity PP-antiserum. Bombesin was infused in a dose of 150 ng/kg·hr, resulting in a rise of plasma PP from 24±5 to 224±25 pM (p<0.01). Before this bombesin infusion, 7 ml of normal rabbit serum had been administered to the dogs (n=8). At a later stage, the study was repeated after administration of 7 ml of PP-antiserum to the same animals. The bombesin induced increase in pancreatic exocrine secretion during administration of PP-antiserum (flow rate 24±10 ml/hr, protein output 1.35±0.43 g/hr, and bicarbonate output 3.25±1.42 mmol/hr) was not significantly different from that during control rabbit serum (flow rate 21±7 ml/hr, protein output 1.26±0.38 g/hr, and bicarbonate output 3.18±1.10 mmol/hr). It is therefore concluded that the pancreatic exocrine response to bombesin is not affected by the concomitant secretion of PP.     

Three-dimensional analysis demonstrates the presence of exocrine cytoplasmic vela between endocrine cells and basal lamina in the stomach of mammals

Three-dimensional analysis demonstrated the presence of cytoplasmic vela extending from exocrine cells into the space between endocrine cells and basal lamina in the gastrointestinal epithelium of the rabbit; these structures were also observed in various other mammals. The following techniques were used to determine the morphologic characteristics of these vela and to study their significance: preparation of semiserial thin sections, three-dimensional reconstruction in plexiglass and lanthanum staining of pericellular spaces. It was found that these fine vela, devoid of major differentiated cell-constituents, sometimes form a pseudocircular crown at the base of endocrine cells. If the zone of basal apposition of the plasma membrane is referred to as ZBA and the zones of lateral apposition as ZLA, the presence of this velum makes it possible to distinguish a zone of immediate apposition without interposition (ZIA) and a mediate zone of apposition with interposition (ZMA) within the ZBA. Exocrine cell processes can also penetrate within endocrine cells in invaginations, and the depth of these invaginations can be demonstrated by lanthanum staining. Adjacent to the membrane zones defined above, other cytoplasmic microdomains-M(ZLA) and M(ZBA), as well as M(ZIA) and M(ZMA) of different morphofunctional significance may also be envisaged.     

Functional anatomy of scent glands in Paranemastoma quadripunctatum (Opiliones,Dyspnoi, Nemastomatidae)

The morphological organization and functional anatomy of prosomal defensive (scent) glands in Paranemastoma quadripunctatum, a representative of the dyspnoid harvestmen, was investigated by means of histological semithin sections, software‐based 3D‐reconstruction and scanning electron microscopy. Scent glands comprise large, hollow sacs on either side of the prosoma, each of these opening to the outside via one orifice (ozopore) immediately above coxa I. In contrast to the situation known from laniatorean, cyphophthalmid and some eupnoid Opiliones, ozopores are not exposed but hidden in a depression (atrium), formed by a dorsal integumental fold of the carapace and the dorsal parts of coxae I. Glandular sacs are connected to ozopores via a short duct which is equipped with a specific closing mechanism in its distal part: A layer of modified epidermal cells forms a kind of pad‐like tissue, surrounding the duct like a valve. Several muscles attached to the anterior parts of the glandular reservoir and to the epithelial pad may be associated with ozopore‐opening. The actual mechanism of secretion discharge seems to be highly unusual and may be hypothesized on the basis of corroborating data from behavioral observations, scent gland anatomy and secretion chemistry as follows: Enteric fluid is considered to be directed towards the ozopores via cuticular grooves in the surface of the coxapophyses of legs I. Then, the fluid is sucked into the anterior part of the scent gland reservoirs by the action of dorsal dilator muscles that widen the reservoir and produce a short‐term negative pressure. After dilution/solution of the naphthoquinone‐rich scent gland contents, a secretion‐loaded fluid is thought to be discharged with the help of transversal compressor muscles. This is the first detailed study on the functional anatomy of scent glands and the mechanisms of secretion discharge in the Dyspnoi. J. Morphol. 2011. © 2011 Wiley‐Liss, Inc.     

Proton microprobe analysis of 15 elements in pancreatic B cells and exocrine pancreas in diabetic Chinese hamsters

Diabetes mellitus spontaneously develops in certain sublines of non-obese Chinese hamsters, and the diabetic L-subline is known for subnormal pancreatic insulin releasein vitro. The cause of the secretory defect is unknown.Freeze-dried pancreas sections from genetically diabetic Chinese hamsters and normal controls were subjected to proton bombardment and the concentration of 15 elements in B cells and acini was calculated from the X-rays emitted. Diabetic B cells contained significantly less Al (–61%) and significantly more Cu (+92 %), Mg (+6 %) and Rb (+13 %) than their normal counterparts. The diabetic acini showed similar, significant changes. The molar ratio between K and Na was about 10 in endocrine as well as exocrine pancreas from both groups of animals, implying that neither sample preparation nor irradiation had induced significant diffusive changes.In conclusion, the high K/Na ratio suggests that the diabetic B cell has a well-functioning Na⁺/K⁺ pump. However, significant and parallel changes in Al-, Cu-, Mg- and Rb-levels were found in both the B cells and acinar portion of the diabetic pancreas. It is not clear whether these elemental changes cause the islet secretory defect or result from it.