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Activation of Ca(2+)-dependent K(+) channels contributes to rhythmic firing of action potentials in mouse pancreatic beta cells.

Göpel SO, Kanno T, Barg S, Eliasson L, Galvanovskis J, Renström E, Rorsman P - J. Gen. Physiol. (1999)

Bottom Line: The current was dependent on Ca(2+) influx but unaffected by apamin and charybdotoxin, two blockers of Ca(2+)-activated K(+) channels, and was insensitive to tolbutamide (a blocker of ATP-regulated K(+) channels) but partially (>60%) blocked by high (10-20 mM) concentrations of tetraethylammonium.This is similar to the interval between two successive bursts of action potentials.We propose that this Ca(2+)-activated K(+) current plays an important role in the generation of oscillatory electrical activity in the beta cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiological Sciences, Division of Molecular and Cellular Physiology, Lund University, SE-223 62 Lund, Sweden.

ABSTRACT
We have applied the perforated patch whole-cell technique to beta cells within intact pancreatic islets to identify the current underlying the glucose-induced rhythmic firing of action potentials. Trains of depolarizations (to simulate glucose-induced electrical activity) resulted in the gradual (time constant: 2.3 s) development of a small (<0.8 nS) K(+) conductance. The current was dependent on Ca(2+) influx but unaffected by apamin and charybdotoxin, two blockers of Ca(2+)-activated K(+) channels, and was insensitive to tolbutamide (a blocker of ATP-regulated K(+) channels) but partially (>60%) blocked by high (10-20 mM) concentrations of tetraethylammonium. Upon cessation of electrical stimulation, the current deactivated exponentially with a time constant of 6.5 s. This is similar to the interval between two successive bursts of action potentials. We propose that this Ca(2+)-activated K(+) current plays an important role in the generation of oscillatory electrical activity in the beta cell.

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Association between [Ca2+]i and Kslow current activation. (A) Membrane current (top) elicited by a train of depolarizations (bottom) and the associated changes of the cytoplasmic [Ca2+]i in an isolated cell (white trace superimposed on current trace). The horizontal line indicate steady state current and [Ca2+]i. (B) The current amplitude measured at the end of the train in isolated cells and in β cells within intact islets. *P < 0.001. (C) Kslow current (top) elicited by the train of depolarizations (bottom) under control conditions and in the presence of 200 μM Cd2+. (D) Voltage-gated Ca2+ currents recorded during 100-ms depolarizations to 0 mV from a holding potential of −70 mV in β cells in intact islets (top) and in dispersed β cells (middle). (E) Charge entry normalized to cell capacitance (Q/C)–voltage (V) relationships of the Ca2+ current recorded in isolated β cells (▴) and in β cells within intact islets (▪). Mean values ± SEM of 8 (▪) and 15 (▴) experiments. *P < 0.05.
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Figure 5: Association between [Ca2+]i and Kslow current activation. (A) Membrane current (top) elicited by a train of depolarizations (bottom) and the associated changes of the cytoplasmic [Ca2+]i in an isolated cell (white trace superimposed on current trace). The horizontal line indicate steady state current and [Ca2+]i. (B) The current amplitude measured at the end of the train in isolated cells and in β cells within intact islets. *P < 0.001. (C) Kslow current (top) elicited by the train of depolarizations (bottom) under control conditions and in the presence of 200 μM Cd2+. (D) Voltage-gated Ca2+ currents recorded during 100-ms depolarizations to 0 mV from a holding potential of −70 mV in β cells in intact islets (top) and in dispersed β cells (middle). (E) Charge entry normalized to cell capacitance (Q/C)–voltage (V) relationships of the Ca2+ current recorded in isolated β cells (▴) and in β cells within intact islets (▪). Mean values ± SEM of 8 (▪) and 15 (▴) experiments. *P < 0.05.

Mentions: Unless otherwise indicated, the electrophysiological experiments were carried out on β cells in intact islets. NMRI mice were purchased from a commercial breeder (Moellegaard). The mice were stunned by a blow against the head and killed by cervical dislocation and the pancreas quickly removed. Collagenase (2 mg) was dissolved in Hank's buffer and injected into the pancreatic duct. Pancreatic islets were isolated by gentle collagenase digestion (25 min, 37°C). Islets thus isolated were subsequently maintained in short-term tissue culture (<16 h) in RPMI 1640 containing 5 mM glucose and 10% (vol/vol) fetal calf serum (Flow Laboratories) and supplemented with 100 μg/ml streptomycin and 100 IU/ml penicillin (both from Northumbria Biologicals, Ltd.). The experiments in Fig. 1C and some of those displayed in Fig. 5 were carried out on dispersed β cells. These were prepared by shaking islets in Ca2+-free solution. The resultant cell suspension was plated on glass cover slips (diameter: 22 mm) or Nunc plastic petri dishes and maintained in tissue culture for up to 48 h using the tissue culture medium mentioned above.


Activation of Ca(2+)-dependent K(+) channels contributes to rhythmic firing of action potentials in mouse pancreatic beta cells.

Göpel SO, Kanno T, Barg S, Eliasson L, Galvanovskis J, Renström E, Rorsman P - J. Gen. Physiol. (1999)

Association between [Ca2+]i and Kslow current activation. (A) Membrane current (top) elicited by a train of depolarizations (bottom) and the associated changes of the cytoplasmic [Ca2+]i in an isolated cell (white trace superimposed on current trace). The horizontal line indicate steady state current and [Ca2+]i. (B) The current amplitude measured at the end of the train in isolated cells and in β cells within intact islets. *P < 0.001. (C) Kslow current (top) elicited by the train of depolarizations (bottom) under control conditions and in the presence of 200 μM Cd2+. (D) Voltage-gated Ca2+ currents recorded during 100-ms depolarizations to 0 mV from a holding potential of −70 mV in β cells in intact islets (top) and in dispersed β cells (middle). (E) Charge entry normalized to cell capacitance (Q/C)–voltage (V) relationships of the Ca2+ current recorded in isolated β cells (▴) and in β cells within intact islets (▪). Mean values ± SEM of 8 (▪) and 15 (▴) experiments. *P < 0.05.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2230648&req=5

Figure 5: Association between [Ca2+]i and Kslow current activation. (A) Membrane current (top) elicited by a train of depolarizations (bottom) and the associated changes of the cytoplasmic [Ca2+]i in an isolated cell (white trace superimposed on current trace). The horizontal line indicate steady state current and [Ca2+]i. (B) The current amplitude measured at the end of the train in isolated cells and in β cells within intact islets. *P < 0.001. (C) Kslow current (top) elicited by the train of depolarizations (bottom) under control conditions and in the presence of 200 μM Cd2+. (D) Voltage-gated Ca2+ currents recorded during 100-ms depolarizations to 0 mV from a holding potential of −70 mV in β cells in intact islets (top) and in dispersed β cells (middle). (E) Charge entry normalized to cell capacitance (Q/C)–voltage (V) relationships of the Ca2+ current recorded in isolated β cells (▴) and in β cells within intact islets (▪). Mean values ± SEM of 8 (▪) and 15 (▴) experiments. *P < 0.05.
Mentions: Unless otherwise indicated, the electrophysiological experiments were carried out on β cells in intact islets. NMRI mice were purchased from a commercial breeder (Moellegaard). The mice were stunned by a blow against the head and killed by cervical dislocation and the pancreas quickly removed. Collagenase (2 mg) was dissolved in Hank's buffer and injected into the pancreatic duct. Pancreatic islets were isolated by gentle collagenase digestion (25 min, 37°C). Islets thus isolated were subsequently maintained in short-term tissue culture (<16 h) in RPMI 1640 containing 5 mM glucose and 10% (vol/vol) fetal calf serum (Flow Laboratories) and supplemented with 100 μg/ml streptomycin and 100 IU/ml penicillin (both from Northumbria Biologicals, Ltd.). The experiments in Fig. 1C and some of those displayed in Fig. 5 were carried out on dispersed β cells. These were prepared by shaking islets in Ca2+-free solution. The resultant cell suspension was plated on glass cover slips (diameter: 22 mm) or Nunc plastic petri dishes and maintained in tissue culture for up to 48 h using the tissue culture medium mentioned above.

Bottom Line: The current was dependent on Ca(2+) influx but unaffected by apamin and charybdotoxin, two blockers of Ca(2+)-activated K(+) channels, and was insensitive to tolbutamide (a blocker of ATP-regulated K(+) channels) but partially (>60%) blocked by high (10-20 mM) concentrations of tetraethylammonium.This is similar to the interval between two successive bursts of action potentials.We propose that this Ca(2+)-activated K(+) current plays an important role in the generation of oscillatory electrical activity in the beta cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiological Sciences, Division of Molecular and Cellular Physiology, Lund University, SE-223 62 Lund, Sweden.

ABSTRACT
We have applied the perforated patch whole-cell technique to beta cells within intact pancreatic islets to identify the current underlying the glucose-induced rhythmic firing of action potentials. Trains of depolarizations (to simulate glucose-induced electrical activity) resulted in the gradual (time constant: 2.3 s) development of a small (<0.8 nS) K(+) conductance. The current was dependent on Ca(2+) influx but unaffected by apamin and charybdotoxin, two blockers of Ca(2+)-activated K(+) channels, and was insensitive to tolbutamide (a blocker of ATP-regulated K(+) channels) but partially (>60%) blocked by high (10-20 mM) concentrations of tetraethylammonium. Upon cessation of electrical stimulation, the current deactivated exponentially with a time constant of 6.5 s. This is similar to the interval between two successive bursts of action potentials. We propose that this Ca(2+)-activated K(+) current plays an important role in the generation of oscillatory electrical activity in the beta cell.

Show MeSH
Related in: MedlinePlus