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  • RINm5F cell  (6)
  • Acinar cells  (5)
  • Membrane potential  (5)
  • 1
    ISSN: 1432-1424
    Keywords: K+ channel ; ATP ; glyceraldehyde ; RINm5F cell
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary The control of K+ channels in the insulin-secreting cell line RINm5F has been investigated by patch-clamp singlechannel current recording experiments. The unitary current events recorded from cell-attached patches are due to large and small inwardly rectifying ATP-sensitive K+ channels with conductance properties similar to the two channels previously identified in primary cultured rat islet cells (Findlay, I., Dunne, M.J., & Petersen, O. H.J. Membrane Biol. 88:165–172, 1985). Cell permeabilization through brief exposure to 10 μm digitonin or 0.05% saponin (outside the isolated membrane patch area) results in a dramatic increase in current through the cell-attached patch due to opening of many large and small K+-selective channels. These channels are inhibited in a dose-dependent manner by ATP applied to the bath (near-complete inhibition by 5mm ATP). During prolonged ATP exposure (1–5 min) the initial inhibition is followed by partial recovery of channel activity, although further activation does occur when ATP is subsequently removed. From the maximal number of coincident channel openings in the permeabilized cells (in the absence of ATP), it is estimated that there are on average 12 large ATP-sensitive K+ channels per membrane patch, but in the intact cells less than 5% of the membrane patches exhibited three or more coincident K+ channel openings, indicating the degree to which the channels are inhibited in the resting condition by endogenous ATP. Stimulation of RINm5F cells to secrete insulin was carried out by challenging intact cells with 10mm d-glyceraldehyde.d-glyceraldehyde induced depolarization of the membrane from about −70 to −20 mV and evoked a marked reduction in the open-state probability of both the large and small ATP-sensitive channels.d-glyceraldehyde also induced action potentials in a number of cases. All effects of stimulation were largely transient, lasting about 100 sec. The two ATP-sensitive K+ channels are probably responsible for the resting potential and play a crucial role in coupling metabolism to membrane depolarization.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-2013
    Keywords: Pancreatic acinus ; Calcium ; Membrane potential ; Membrane resistance ; EGTA ; Stimulus-permeability coupling
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Membrane potential and resistance were measured from two neighbouring electrically coupled cells within the same mouse pancreatic acinus, using one electrode filled with KCl and another filled with K-EGTA [ethylene glycol-bis-(β-amino ethyl ether) N,N′-tetra acetic acid] or K-acetate. EGTA was injected into one cell by passing pulses of hyperpolarizing current through the EGTA microelectrode. In order to ensure constant membrane potential, pulses of depolarizing current of the same intensity were simultaneously passed through the other electrode. Intracellular injection of EGTA, but not acetate, decreased significantly the membrane response (depolarization, resistance reduction) of both coupled cells to small microionophoretic pulses of ACh stimulation. Responses to larger doses were decreased to a smaller extent. Intracellular injection of EGTA also decreased significantly the membrane response of both coupled cells to microionophoretic pulses of pentagastrin stimulation. In contrast intracellular injection of EGTA did not inhibit membrane response tol-alanine stimulation.l-Alanine-evoked membrane depolarization was independent of external Ca. This is in contrast to what has previously been shown for secretagogue effects. It is concluded that EGTA-induced sequestration of cytosolic Ca2+ decreases membrane responses to both ACh and pentagastrin, providing new evidence of the crucial role played by internal Ca2+ in mediating stimulus-permeability coupling. In contrast,l-alanine action does not appear to be mediated by internal Ca2+.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 386 (1980), S. 153-159 
    ISSN: 1432-2013
    Keywords: l-Amino acids ; Membrane potential ; Membrane resistance ; Pancreatic acinar cell ; Na-conductance ; l-Alanine-amino acid interactions
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The effect of some neutrall-amino acids, alanine, valine and proline, on the pancreatic acinar cell membrane potential and resistance was investigated. Simultaneous recordings were made with two intracellular microelectrodes on isolated superfused segments of mouse pancreas. Amino acids were applied by inclusion at known concentrations in the superfusion fluid or by microionophoresis from extracellular micropipettes. l-Alanine (10 mmol·l−1) evoked a maximal membrane depolarization of about 18 mV. A just detectable depolarization was observed at 0.1 mmol·l−1 (3 mV). Halfmaximal depolarization was observed at 1.6 mmol·l−1.d-Alanine had virtually no effect. Microionophoretic applications ofl-alanine,l-valine orl-proline evoked depolarization and resistance reduction with a very short delay (〈50 ms). The dose response curves for depolarization and resistance reduction were similar. The amplitude of the depolarization evoked byl-alanine,l-valine andl-proline depended linearly on the level of the pre-set membrane potential (membrane potential could be changed by direct current injection). With decreasing intracellular negativity there was a decrease in the size of the amino acid-evoked depolarization. When the membrane potential was inside positive the amplitude became very small. Extrapolation of the linear relations between membrane potential and size of depolarization revealed a null potential of +20 to +45 mV. Thel-alanine-evoked depolarization was acutely reduced but not abolished by replacing extracellular Na by Tris or Li. l-Alanine,l-proline andl-valine exhibited mutual inhibition of evoked depolarization even when the depolarizing effect of the first applied amino acid was balanced by direct current injection. It is concluded that severall-amino acids act on the pancreatic acinar plasma membrane by opening conductance pathways mainly permeable to Na.
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 400 (1984), S. 51-59 
    ISSN: 1432-2013
    Keywords: Lacrimal acinar cell ; Membrane potential ; Membrane resistance ; Nerve stimulation ; Electrical field stimulation ; Acetylcholine ; Adrenaline
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Intracellular microelectrode recordings were made from superfused in vitro preparations of mouse lacrimal giand. The lacrimal acinar cell had a mean resting membrane potential of −44.1±0.5 mV and a mean input resistance of 3.5±0.15 MΩ. Electrical field stimulation (FS) had similar effects to ACh applied by microionophoresis, both evoking a biphasic membrane hyperpolarization (up to 15 mV) accompanied by a reduction in input resistance. The equilibrium potential values (EFS and EACh) for the responses to brief duration FS and ACh ionophoresis ranged between −45 and −75 mV and depended on the time at which measurements were made following the onset of stimulation. Superfusion of ACh or adrenaline also caused membrane hyperpolarization and increased membrane conductance. Estimations of EFS and EACh made during prolonged periods of FS and ACh superfusion yielded mean values of −53.9±1.9 mV and −53.4±1.5 mV respectively. FS evoked a response in all preparations tested with maximal effects seen at 40 Hz frequency. The mean latency of the FS-evoked hyperpolarization (40 Hz) was 270±21 ms and that for the ACh ionophoretic response was 400±65 ms. Low frequency FS (0.5–5 Hz) also induced membrane hyperpolarization and responses to single shock stimuli were occasionally observed. The FS-evoked hyperpolarization was abolished following the blockade of nerve conduction by superfusion of either Na-free or tetrodotoxin-containing media. Effects of FS were not seen in the presence of atropine. Neostigmine potentiated the FS- and ACh-evoked hyperpolarizations. Spontaneous miniature hyperpolarizations were unaffected by tetrodotoxin but abolished by atropine. It is concluded that FS excites a well developed cholinergic innervation of the mouse lacrimal gland resulting in ACh release and acinar cell hyperpolarization. ACh, which appears to be the only neurotransmitter released, mediates its effects by increasing plasma membrane permeability to mainly K ions.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1432-2013
    Keywords: K+ channels ; Acinar cells ; Ensemble noise analysis ; Current relaxation ; Patch-clamp whole cell recording
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The K+ channel in rat parotid gland acinar cells were investigated by ensemble current noise analysis in single isolated cells employing the giga-seal whole cell current recording mode. Sets of 20–40 identical de- and hyperpolarization voltage steps were applied and the resultant current records were processed by computer to obtain the mean and the variance of the current. The time-course of the mean current could be fitted by the sum of two exponentials, suggesting a 3-state model. The simplest plausible hypothesis is a model with one open and two closed states. Assuming this model, the relationship between the variance (σ2) and the mean current (I) could be fitted by the function σ2/I=i−I/N. The estimated single channeli/V-relations were similar to those taken from single channel current recordings, and the size of the population of channels per cell (N) was 76±26 (n=12). The validity of the model was tested by a successful simulation of the time-course of the variance.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1432-2013
    Keywords: Patch-clamp single channel recording ; K+ channel ; K+ conductance ; Rb+ conductance ; Salivary gland ; Pancreas ; Acinar cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Single-channel current recordings were carried out on excised inside-out patches of baso-lateral plasma membrane from exocrine acinar cells. The mouse pancreas and submandibular gland as well as the pig pancreas were investigated. In the mouse pancreas the voltage-insensitive Ca2+-activated cation channel was studied. Single-channel current-voltage (i/v) relationships were studied in symmetrical Rb+-rich solutions and in asymmetrical Rb+/Na+ and Na+/Rb+ solutions. In all cases the i/v relations were linear and had the same slope representing a single-channel conductance of about 33 pS which is identical to that previously obtained with symmetrical Na+ solutions or asymmetrical Na+/K+ solutions. In the mouse submandibular gland and the pig pancreas the voltage and Ca2+-activated K+ channel was studied. The outward currents observed after depolarization in the presence of quasi-physiological Na+/K+ gradients were immediately abolished when all the K+ in the bath fluid was replaced by Rb+ (bath fluid in contact with inside of plasma membrane). This effect was immediately and fully reversible upon return to the high K+ solution. The voltage and Ca2+-activated K+ channel was also studied in asymmetrical K+/Rb+ and Rb+/K+ solutions. In the first case inward (K+) currents could be observed but not outward (Rb+) currents, while in the other case inward (Rb+) currents could not be seen whereas outward (K+) currents were measured. The current-voltage relationships were approximately linear and the null potential was close to 0 mV in both situations. In contrast the null potential for current through the K+ channel in the presence of asymmetrical Na+/K+ or Li+/K+ solutions was about −70 mV and with reversed gradients about +60 mV. Outward K+ currents of reduced size (through the voltage and Ca2+-activated K+ channel) could be observed when the bath fluid contained 75 mM K+ and 75 mM Rb+, but not (in the same membrane patches) when 150 mM Rb+ and no K+ was present. It is concluded that the large voltage- and Ca2+-activated K+ channel has an extremely low Rb+ conductance. It is possible, however, that the permeability for Rb+ may be about the same as for K+. The voltage-insensitive Ca2+-activated cation channel does not discriminate between K+ and Rb+.
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  • 7
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 391 (1981), S. 119-124 
    ISSN: 1432-2013
    Keywords: Thyroid gland ; Follicular cell ; Membrane potential ; Membrane resistance ; Cell to cell coupling ; Specific membrane resistance ; Electrogenic pump
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract Intracellular recordings of membrane potential, input resistance and time constant have been made in vitro from the follicular cells of the rat, rabbit and guinea-pig thyroid glands using glass microelectrodes. The passive permeability properties of these cells have been investigated by altering the concentration of one or more ions in the superfusing fluid. Investigations into the intercellular coupling characteristics of the thyroid gland were made by inserting two microelectrodes into neighbouring communicating cells. The mean transmembrane potentials were between −60 and −70 mV in all three species studied. The magnitude of the membrane potential in the rat was found to be dependent mainly upon the gradient for potassium (K+) across the membrane. Current-voltage relationships were investigated in all three species by injecting rectangular de- or hyperpolarizing current pulses through the recording microelectrode. Within a relatively wide range (−20 to −80 mV), there was an approximately linear relationship between injected current and change in membrane potential. The input resistance was about 11 MΩ in all three species, while the time constant (τ) varied from 5–35 ms. Readmitting K to K-deprived rat thyroids during intracellular microelectrode recording caused a transient hyperpolarization which was unaccomapanied by any change in input resistance. The transient hyperpolarization was abolished by ouabain. Addition of 10−3 M ouabain to the resting cell caused an immediate depolarization of approximately 2 mV. Electrical coupling between neighbouring cells could only be observed if the distance between the tips of the two exploring microelectrodes was less than 15 μm. The coupling coefficient (V 2/V 1) was close to 1. Assuming uniform current spread within one follicle and electrical isolation of individual follicles from each other the specific membrane resistance of the rat thyroid follicular cells was calculated to be 4.9 kΩcm2.
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  • 8
    Electronic Resource
    Electronic Resource
    Springer
    Pflügers Archiv 382 (1979), S. 263-267 
    ISSN: 1432-2013
    Keywords: Pancreatic acinar cells ; N2, O2′ dibutyryl guanosine 3′:5′-Monophosphate ; Cholecystokinin antagonist ; Bombesin ; Membrane potential
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract The effects of dibutyryl cyclic guanosine 3′:5′ monophosphate (dbcGMP) on the electrical responses of in vitro mouse pancreatic acinar cells to caerulein, bombesin and acetylcholine, applied by microionophoresis, were investigated using intracellular microelectrode recordings. Exposure to dbcGMP (10−3 mol ·l−1) quickly abolished the depolarization response to caerulein ionophoresis, while the bombesin-nonapeptide and acetylcholine-induced depolarizations were retained. Exposing acinar cells to 2×10−4 mol·l−1 dbcGMP reduced their sensitivity such that a 10-fold increase in the applied caerulein ionophoresis current was required to restore the responses to the same magnitude as those obtained in a dbcGMP-free environment. The response to topical applications of pentagastrin and CCK-33 were also abolished by dbcGMP. The results provide direct evidence that functional ACh, bombesin and CCK receptors are all present within the same acinar cell unit. The dbcGMP-induced inhibition of responses evoked by CCK-like peptides is immediate, specific and easily reversible. DbcGMP acts as a competitive antagonist of the CCK receptor on pancreatic acinar cells.
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  • 9
    ISSN: 1432-2013
    Keywords: Ca2+ extrusion ; Ca2+ measurements ; Droplet technique ; Acinar cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Medicine
    Notes: Abstract This paper contains a description of the modified droplet technique that is designed to monitor Ca2+ extrusion from single isolated pancreatic acinar cells. A cell loaded with calcium indicator is maintained in a small droplet of solution containing another calcium indicator. Differences in the optical properties of the intracellular and extracellular indicators allows one to monitor simultaneously intracellular and extracellular calcium concentrations. The paper contains a description of the calibration procedure that is used to calculate intracellular and extracellular calcium concentrations. The advantages and disadvantages of different pairs of extracellular and intracellular indicators are discussed.
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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    The journal of membrane biology 102 (1988), S. 205-216 
    ISSN: 1432-1424
    Keywords: K+ channel ; ATP ; NAD(P) ; NAD(P)H ; RINm5F cell
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Summary The single-channel current recording technique has been used to study the influences that the pyridine nucleotides NAD, NADH, NADP and NADPH have on the gating of ATP-sensitive K+ channels in an insulin-secreting cell line (RINm5F). The effects of the nucleotides were studied at the intracellular surface using either excised inside-out membrane patches or permeabilized cells. All four pyridine nucleotides were found to evoke similar effects. At low concentrations, 100 μm and less, each promoted channel opening whereas high concentrations, 500 μm and above, evoked channel closure. The degree of K+ channel activation by pyridine nucleotides (low conc.) was found to be similar to that evoked by the same concentrations of ADP or GTP, whereas the degree of K+ channel inhibition (high conc.) was less marked than that evoked by the same concentrations of ATP, and never resulted in refreshment of K+ channels following removal. The effects of NAD, NADH, NADP and NADPH seemed to interact with those of ATP and ADP. In the presence of 1mm ADP and 4mm ATP, 10 to 100 μm concentrations of the pyridine nucleotides could not evoke channel opening, whereas concentrations of 500 μm and above were found to evoke channel closure. In the presence of 2mm ATP and 0.5mm ADP, however, 10 to 100 μm concentrations of the pyridine nucleotides were able to activate K+ channels.
    Type of Medium: Electronic Resource
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