Age-dependent changes in the exocytotic efficacy in Kir6.2 ablated mouse pancreatic beta cells

by Ernest Beaudelaire Tsiaze
Doctoral thesis
Date of Examination:2014-04-02
Date of issue:2014-03-31
Advisor:Prof. Dr. Marjan Rupnik
Referee:Prof. Dr. Tobias Moser
Referee:Prof. Dr. Blanche Schwappach
Persistent Address: http://dx.doi.org/10.53846/goediss-4437

 

 

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Abstract

English

ATP-sensitive K+ channels (K channels) regulate cellular energy metabolism in control of membrane excitability. The KATP channel of pancreatic β-cells is composed of SUR1 and Kir6.2 subunits. Kir6.2-/- mice have a defective KATP channel and can mimic the phenotype of persistent hyperinsulinaemia hypoglycaemia of infancy (PHHI) where the stimulus-secretion coupling underlying insulin secretion is disrupted. In this study, a transient hypoglycaemia in young (2 - 4 weeks old) and a progressive hyperglycaemia in ageing (5 - 60 weeks old) Kir6.2-/- mice were observed compared to control. Furthermore, a pancreatic tissue slice preparation was used as a novel technique to reassess islets morphology and to study the electrophysiological properties of Kir6.2-/- β-cells in a near native environment. Using this novel approach, age-linked differences in cyto-architecture reflecting the defective glucose homeostasis were seen in Kir6.2-/- mice. Moreover, the coupling of membrane excitation to exocytosis was verified using the patch clamp technique that enables a high resolution measurement of membrane capacitance (Cm) changes upon exocytosis of hormone. I observed a more efficient stimulus-secretion coupling in β-cells of young Kir6.2-/- mice compared to ageing Kir6.2-/- mice. This enhanced stimulus-secretion coupling was further verified by the application of repetitive trains of stimulation. To test if the more efficient stimulus-secretion coupling was due to enhanced calcium (Ca2+) sensitivity of exocytosis, the efficacy of Ca2+ to trigger secretion was analyzed and found to be increased. Additionally, the constitutive bursting and spiking electrical activity as well as the larger high voltage activated (HVA) Ca2+current density monitored in pancreatic β-cells from Kir6.2-/- mice may explain the known elevated basal [Ca2+]c and provide more insight in clarifying the late steps of stimulus-secretion coupling during glucose metabolism in Kir6.2-/- β-cells. These findings should also promote the understanding of the pathophysiology of PHHI and other KATP channels related pancreatic diseases.
Keywords: KATP channels; Kir6.2 ablated mice; Persistent Hyperinsulinaemia Hypoglycaemia of Infancy; Pancreatic tissue slice; Patch-clamping; Exocytosis
 
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