Pharmacological stimulation and inhibition of insulin secretion in mouse islets lacking ATP-sensitive K+ channels

Abstract

Background and purpose: ATP-sensitive potassium channels (K(ATP) channels) in beta cells are a major target for insulinotropic drugs. Here, we studied the effects of selected stimulatory and inhibitory pharmacological agents in islets lacking K(ATP) channels.

Experimental approach: We compared insulin secretion (IS) and cytosolic calcium ([Ca(2+)](c)) changes in islets isolated from control mice and mice lacking sulphonylurea receptor1 (SUR1), and thus K(ATP) channels in their beta cells (Sur1KO).

Key results: While similarly increasing [Ca(2+)](c) and IS in controls, agents binding to site A (tolbutamide) or site B (meglitinide) of SUR1 were ineffective in Sur1KO islets. Of two non-selective blockers of potassium channels, quinine was inactive, whereas tetraethylammonium was more active in Sur1KO compared with control islets. Phentolamine, efaroxan and alinidine, three imidazolines binding to K(IR)6.2 (pore of K(ATP) channels), stimulated control islets, but only phentolamine retained weaker stimulatory effects on [Ca(2+)](c) and IS in Sur1KO islets. Neither K(ATP) channel opener (diazoxide, pinacidil) inhibited Sur1KO islets. Calcium channel blockers (nimodipine, verapamil) or diphenylhydantoin decreased [Ca(2+)](c) and IS in both types of islets, verapamil and diphenylhydantoin being more efficient in Sur1KO islets. Activation of alpha(2)-adrenoceptors or dopamine receptors strongly inhibited IS while partially (clonidine > dopamine) lowering [Ca(2+)](c) (control > Sur1KO islets).

Conclusions and implications: Those drugs retaining effects on IS in islets lacking K(ATP) channels, also affected [Ca(2+)](c), indicating actions on other ionic channels. The greater effects of some inhibitors in Sur1KO than in control islets might be relevant to medical treatment of congenital hyperinsulinism caused by inactivating mutations of K(ATP) channels.

Figure 1

Effects of tolbutamide (100 µmol·L⁻¹) and diazoxide (100 µmol·L⁻¹) on [Ca²⁺]_c (panel A) and insulin secretion (panel B) in control islets perifused with a medium containing 15 mmol·L⁻¹ glucose throughout. Values are means ± SE for 16–30 islets from 3–5 preparations in [Ca²⁺]_c experiments, and 5–12 perifusions in insulin secretion experiments. The figure illustrates how the effects of the drugs were computed for presentation in tables. In each individual experiment, [Ca²⁺]_c and the insulin secretion rate were averaged between 5 and 15 min to obtain the reference value (100%). Steady-state effects of drugs were averaged between 30 and 40 min for [Ca²⁺]_c, and 30–45 min for insulin secretion, and expressed relative to the reference value in the same experiment. The mean effect of the drug was then calculated. Panel A thus shows that [Ca²⁺]_c increased by 13% in the absence of drug and by 67% after addition of tolbutamide, and that it decreased by 75% after addition of diazoxide.

Figure 2

Effects of selected stimulators of insulin secretion on [Ca²⁺]_c in control and Sur1KO islets perifused with 15 mmol·L⁻¹ glucose. Meglitinide (100 µmol·L⁻¹), tetraethylammonium (TEA, 10 mmol·L⁻¹), phentolamine (100 µmol·L⁻¹) or alinidine (100 µmol·L⁻¹) was added as indicated at the top of each panel. Traces illustrate the changes occurring in representative islets. Quantification of the changes is presented in Table 1.

Figure 3

Effects of selected inhibitors of insulin secretion on [Ca²⁺]_c in control and Sur1KO islets perifused with 15 mmol·L⁻¹ glucose. Pinacidil (250 µmol·L⁻¹), verapamil (20 µmol·L⁻¹), diphenylhydantoin (DPH, 20 µmol·L⁻¹) or clonidine (1 µmol·L⁻¹) was added as indicated at the top of each panel. Traces illustrate the changes occurring in representative islets. Quantification of the changes is presented in Table 2.