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  • 1
    ISSN: 1520-4995
    Source: ACS Legacy Archives
    Topics: Biology , Chemistry and Pharmacology
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-2013
    Keywords: Ca2+-activated K+ channels ; Maxi-K+ channel ; Tetraethylammonium ; Salivary gland ; Fluid secretion ; K+ efflux ; Epithelial transport
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The involvement of large-conductance, voltage- and Ca2+-activated K+ channels (maxi-K+ channels) in basolateral Ca2+-dependent K+-efflux pathways and fluid secretion by the rat submandibular gland was investigated. Basolateral K+ efflux was monitored by measuring the change in K+ concentration in the perfusate collected from the vein of the isolated, perfused rat submandibular gland every 30 s. Under conditions in which the Na+/K+-ATPase and Na+-K+-2Cl− cotransporter were inhibited by ouabain (1 mmol/l) and bumeta-nide (50 μmol/l) respectively, continuous stimulation with acetylcholine (ACh) (1 μmol/l) caused a transient large net K+ efflux, followed by a smaller K+ efflux, which gradually returned to the basal level within 10 min. These two components of the K+ efflux appear to be dependent on an increase in cytosolic Ca2+ concentration. The initial transient K+ efflux was not affected by charybdotoxin (100 nmol/l) or tetraethylammonium (TEA) (5 mmol/l) but the smaller second component was strongly and reversibly inhibited by charybdotoxin (100 nmol/l) and TEA (0.1 and 5 mmol/l). The initial K+ efflux transient induced by ACh was inhibited by quinine (0.1–3 mmol/l), quinidine (1–3 mmol/l) and Ba2+ (5 mmol/l), but not by verapamil (0.1 mmol/l), lidocaine (1 mmol/l), 4-aminopyridine (1 mmol/l) or apamin (1 μmol/l). Ca2+-dependent transient large K+ effluxes induced by substance P (0.01 μmol/l) and A23187 (3 μmol/l) were not inhibited by TEA (5 mmol/l or 10 mmol/l). A23187 (3 μmol/l) evoked a biphasic fluid-secretory response, which was not inhibited by TEA (5 mmol/l). Patch-clamp studies confirmed that the whole-cell outward K+ current attributable to maxi-K+ channels obtained from rat submandibular endpiece cells was strongly inhibited by the addition of TEA (1–10 mmol/l) to the bath. It is concluded that maxi-K+ channels are not responsible for the major part of the Ca2+-dependent basolateral K+ efflux and fluid secretion by the rat submandibular gland.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1432-2013
    Keywords: Intracellular Na ; Na influx ; Acetylcholine ; Ouabain ; 23Na NMR ; Salivary gland
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Intracellular Na content (Nain) in the perfused rat mandibular gland was measured by using a23Na NMR spectroscopy at 24°C. An aqueous chemical shift reagent, dysprosium triethylenetetramine-N,N,N′,N″,N‴N‴-hexaacetic acid [Dy(TTHA)] was used in order to discriminate between the intracellular and the extracellular Na signal. The mandibular gland of rat was perfused arterially with a modified Krebs solution containing 10 mM Dy(TTHA). At rest, Nain was not changed by blocking the Na+/K+ ATPase with ouabain (1 mM) and atropine (3 μM), implying that, in the absence of stimulation, the spontaneous Na influx across the plasma membrane must have been negligibly small. Following onset of stimulation with acetylcholine (1 μM), Nain increased by 9.1±1.5 mmol/l intracellular fluid (mean±SEM,n=13), and remained at this level during stimulation. In the initial phase of secretion (0–5 min), about 50 mmol/min/l intracellular fluid of Na was secreted into the luminal space (estimated from the secretory rate by assuming an isotonic primary secretion) but, in spite of the higher secretion rate, Nain increased only at an initial rate of 4.1 mmol/min/l intracellular fluid. During the steady phase of secretion (15–30 min) evoked by acetylcholine (1 μM), ouabain (1 mM) caused an increment of Nain of 44±8 mmol/l intracellular fluid (mean±SEM,n=4). From the rate of Nain increment, the Na influx rate at the steady phase was estimated as 4.5 mmol/min/l intracellular fluid. These results suggest that the influx of Na is caused by stimulation with acetylcholine. The observed Na influx rate was about 50% of the Na secretory rate at the steady phase of secretion, estimated from the secretory rate by assuming an isotonic primary secretion. This is fully compatible with the operation of Na−K-2Cl contransport system for which one would expect a Na influx rate exactly half of the rate of Na and Cl secretion.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1432-2013
    Keywords: Salivary gland ; Intracellular pH ; 31P NMR spectroscopy ; Acetylcholine ; Amiloride ; DIDS ; Na+−H+ exchange
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Intracellular pH (pHi) was measured in the isolated, perfused rabbit mandibular salivary gland by31P NMR spectroscopy. In the unstimulated gland perfused with HCO 3 − /CO2-buffered Ringer's solution, pHi was 7.27±0.01. Continuous stimulation with acetylcholine elicited dose- and time-dependent changes in pHi. 10−6 mol/l acetylcholine caused a brief intracellular acidosis (−0.19±0.06 pH units) followed by an increase in pHi to a more alkaline steady-state value (7.33±0.02). In the absence of perfusate HCO 3 − or in the presence of 10−4 mol/l DIDS (4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid), the transient acidosis was abolished and pHi increased rapidly to give a sustained alkalosis (7.49±0.03 and 7.44±0.03 respectively). In the presence of 10−3 mol/l amiloride, the response to acetylcholine was a rapid decrease in pHi to 7.02±0.02. The data suggest that, during perfusion with HCO 3 − /CO2-buffered solutions, stimulation with acetylcholine results in a transient loss of HCO 3 − from the acinar cells (causing a transient acidosis), and, independently, the activation of Na+−H+ exchange (causing a sustained alkalosis). In the unstimulated gland, DIDS and the HCO 3 − -free perfusate caused decreases in pHi to 7.12±0.02 and 7.04±0.01 respectively. In contrast, amiloride had little effect. The relatively high value of pHi maintained by the unstimulated gland is therefore probably not due to Na+−H+ exchange.
    Type of Medium: Electronic Resource
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