Biomedical subjects

T D Plant

Publications and source records attributed to T D Plant.

24 records · Page 2Linked to original sources

PubMed1988

Na+ currents in cultured mouse pancreatic B-cells.

Pancreatic B-cells, kept in culture for 1-4 days, were studied in the whole-cell, cell-attached and outside-out modes of the patch clamp technique. B-cells were identified by the appearance of electrical activity in the cell-attached mode when the bath glucose was raised from 3 to 20 mM. In whole-cell, 80% of these cells showed a transient inward Na+ current, when depolarizing pulses were preceded by holding potentials, or prepulses to potentials more negative than -80 mV. The midpoint (Eh) of the inactivation curve (h infinity) was at -109 mV in 2.6 mM Ca2+, 1.2 mM Mg2+ and -120 mV in 0.2 mM Ca2+, 3.6 mM Mg2+. In 2.6 mM Ca2+, inactivation was fully removed at E less than -150 mV. Na+ currents activated at E greater than -60 mV and were largest at around -10 mV (120 mM Na+). The kinetic parameters of activation (tp) and inactivation (tau h) were similar to those of other mammalian Na+ channels. Unitary currents with an amplitude of approximately 1 pA at -30 mV (140 mM Na+) with a similar voltage-dependence and time-course of mean current were recorded from outside-out patches. The results show that B-cells have a voltage-dependent Na+ current which, owing to the voltage-dependence of inactivation, is unlikely to play a major role in glucose-induced electrical activity.

Animals↗

PubMed1988

Properties and calcium-dependent inactivation of calcium currents in cultured mouse pancreatic B-cells.

1. Ca2+ currents were recorded using the whole-cell mode of the patch-clamp technique from mouse pancreatic B-cells kept in culture for 1-4 days. B-cells were identified in the cell-attached mode by their response to a change in the glucose concentration from 3 to 15 or 20 mM or by their inward currents. 2. Only one component of Ca2+ current was observed in these cells, which activated at potentials greater than -50 mV and was blocked by nitrendipine (5 microM), and increased in amplitude by CGP 28392 (5 microM). 3. During maintained depolarizations the Ca2+ current inactivated considerably but not completely. Inactivation was most marked at potentials where the Ca2+ currents were large, but in general was slower for currents at potentials greater than 0 mV than at more negative potentials. 4. Two-pulse experiments showed that the inactivation curve for the Ca2+ current was U-shaped, returning to unity at potentials approaching the Ca2+ equilibrium potential. Measurements of Ca2+ entry showed that inactivation was dependent on the amount of Ca2+ entering during the pre-pulse, independent of the pre-pulse potential. 5. Ca2+ currents were not appreciably slowed when BAPTA, a faster buffer of Ca2+, replaced EGTA in the pipette solution. 6. Replacement of Ca2+ in the external solution by Ba2+ increased the amplitude of the inward current and largely abolished inactivation. Large inward currents through Ca2+ channels were observed in the absence of divalent cations in the external solution (+EGTA), which were presumably carried by Na+. These currents did not inactivate during 150 ms depolarizations, but were increased in amplitude by CGP 28392 (5 microM) and blocked by D600 (30 microM). 7. The observations suggest that normal mouse pancreatic B-cells have only one type of Ca2+ channel which is dihydropyridine sensitive and inactivates by a mechanism which is almost purely Ca2+ dependent. Inactivation of the Ca2+ current will probably be important in the control of Ca2+ entry during glucose-induced electrical activity.

Action Potentials↗

PubMed1986

The effects of rubidium ions on components of the potassium conductance in the frog node of Ranvier.

The effects of replacement of external and internal K+ ions by Rb+ ions on the two fast components (gf1 and gf2) and slow component (gs) of the K+ conductance (gK) in frog nodes of Ranvier were investigated under voltage- and current-clamp conditions. Fast and slow components of gK were separated by double exponential fits to tail currents following long depolarizing pre-pulses, or by the use of short pre-pulses which activate little gs X gs was also isolated by 1 mM-4-aminopyridine (4-AP). gf1 and gf2 were distinguished in the fast conductance-voltage curve by their different voltage dependences, gf1 activating at more negative potentials. Reversal potential measurements indicated that Rb+ is less permeant than K+, and measurements in 4-AP indicated that the slow component has a lower Rb+ permeability than the fast. In a 50% K+, 50% Rb+ mixture PRb/PK was less than that in 100% Rb+ suggesting that PRb/PK is mole-fraction dependent. With external Rb+ the current-voltage relation was shifted by ca.-10 mV compared to that in K+, an effect on gf ( = gf1 + gf2). The slow conductance (gs) and, under similar conditions, the Na+ current-voltage relation were not shifted. gf, calculated from inward tail currents, was reduced with external Rb+ at potentials where gf2 was activated. Instantaneous current-voltage relations following pre-pulses which activate different components of gf confirmed these observations. In K+ the instantaneous current-voltage relation showed some inward rectification which was largely abolished with Rb+. Comparison of gf calculated from outward (go) and inward (gi) currents confirmed this, and showed that inward gf2 was reduced with Rb+ such that go = gi. Outward currents were little affected by external Rb+. External Rb+ slowed the fast inward tail current following all pre-pulses which activate gf, but had no effect on the time course of the slow component of the tail current. Regenerative responses, which occur in high [K+] (+300 nM-tetrodotoxin) solutions in current clamp did not repolarize in Rb+. Voltage-clamp experiments showed that inactivation of inward currents is slowed when Rb+ is the charge carrier. Replacement of internal K+, by application of Rb+ to the cut ends of the fibre, shifted the reversal potential to more positive potentials but had no effect on the conductance or kinetics. External Rb+ has a large number of effects on inward currents, but little effect on outward currents. Internal Rb+ had little effect on outward or inward currents.(ABSTRACT TRUNCATED AT 400 WORDS)

4-Aminopyridine↗

PubMed1983

The effects of injection of calcium ions and calcium chelators on calcium channel inactivation in Helix neurones.

1. The effects of intracellular injection of Ca, EGTA and EGTA/Ca buffers on inward currents flowing through the Ca channel in Helix aspersa neurones were studied under voltage clamp. 2. Inward currents were reversibly reduced by Ca injection. The extent of the reduction was dependent on the size and duration of the injection. Recovery from injection was exponential with a time constant around 18 s at 18-20 degrees C. 3. In our salines, which contained tetraethylammonium chloride and 4-aminopyridine, no outward current was activated by Ca injection at the holding potential. A given Ca injection reduced the inward current by the same fraction in 25 mM- and 2 . 5 mM-Sr and also at different test potentials. We conclude that Ca injection does not activate an outward current. 4. Mean resting ionized internal Ca concentration, [Ca]i, measured with a Ca-sensitive electrode was 1.9 X 10(-7) M. Our injections increased this by less than 10(-5) M, as expected if most of the injected Ca is bound. Constant-field or Eyring rate theory considerations suggest that this rise in [Ca]i would not significantly affect the inward current through open Ca channels and we conclude, therefore, that Ca injection causes Ca channel inactivation. 5. The effect of Ca injection was blocked by prior injection of EGTA. Extracellular application of carbonyl cyanide m-chlorophenylhydrazone increased the effect of Ca injection. 6. Ca injection does not inactivate Ca channels by lowering pHi since large H+ injections only caused small irreversible decreases in inward current. 7. EGTA injection increased peak Ca current (ICa) by around 30% and decreased the rate and extent of inactivation. Some inactivation remained, however, even after maximal EGTA injections. 8. Injection of EGTA/Ca buffers with free [Ca] less than 1.8 X 10(-7) M increased peak ICa, while buffers with free [Ca] greater than 8.9 X 10(-7) M decreased ICa. 9. Our results support the suggestion that Ca ions cause Ca inactivation by binding to a site which is accessible from the inside of the cell, and also suggest that there is some steady-state inactivation present at physiological [Ca]i. A simple model is presented which describes the decline of Ca current in terms of Ca binding to a site accessible from the cytoplasm.

Animals↗

PubMed1981

Gating of a muscle K+ channel and its dependence on the permeating ion species.

In excitable cells, ions permeate the cell membrane through ionic channels, some of which open and close in response to changes in the potential difference across the membrane. It has been supposed that this opening and closing (or gating) process is largely independent of the permeating ion. However, we show here that the gating of the resting potassium permeability of frog skeletal muscle depends on the species of ion which carries current across the membrane. The potassium permeability investigated allows K+ to move in across the membrane more easily than out. This property is known as inward or anomalous rectification and is shared by cell membranes of skeletal muscle, egg and certain other cells. In both egg cells and skeletal muscle fibres, the group IIIB metal ion Tl+, which can replace K+ in several other systems in experimental conditions, also permeates the inward rectifier. Indeed, Tl+ is more permeant than K+ (refs 8, 9). However, when Tl+ carries current inwards across the membrane, the inward rectifier inactivates over a brief period when the membrane is hyperpolarized, whereas when K+ carries current, the permeability increases with time under hyperpolarization.

Animals↗

PubMed1981

Calcium current inactivation in identified neurones of Helix aspersa.

1. A two-electrode voltage clamp method was used to study Ca inward currents in identified Helix aspersa neurones bathed in 25 mM-Ca, Na-free saline with TEA and 4-AP. 2. Inward currents were blocked by CdCl2. In Cd delayed outward currents appeared at +30 mV. When two identical depolarizations were separated by a gap inward current turned off to the same level during the two pulses up to +20 mV; above this potential the records cross over. 3. The turn-off of inward current at potentials up to +20 mV was not affected by 0.2 mM-quinine, which reduced outward currents at more depolarized potentials. Inward currents declined exponentially over the first 100 msec with a time constant around 60 msec at 0 mV. Double-pulse experiments gave the same time course of turn-off. 4. When Ca inward current was reduced by lowering [Ca]o or by partial block by Cd the rate and extent of turn-off was reduced. 5. The inactivation curve (obtained using a double pulse with gap method) was U-shaped. The curve was not significantly changed by addition of quinine (0.2 mM) or by changing test pulse size. 6. Recovery of inward currents after a depolarizing prepulse was a double-exponential process, with time constants of 120 msec and 9.4 sec at 10--11 degrees C. 7. Our results are discussed in terms of possible Ca-dependent Ca inactivation and in terms of the possibility of development of an outward Ca-dependent K current.

4-Aminopyridine↗