Progression of diet-induced diabetes in C57BL6J mice involves functional dissociation of Ca2(+) channels from secretory vesicles.
OBJECTIVE
The aim of the study was to elucidate the cellular mechanism underlying the suppression of glucose-induced insulin secretion in mice fed a high-fat diet (HFD) for 15 weeks.


METHODS
C57BL6J mice were fed a HFD or a normal diet (ND) for 3 or 15 weeks. Plasma insulin and glucose levels in vivo were assessed by intraperitoneal glucose tolerance test. Insulin secretion in vitro was studied using static incubations and a perfused pancreas preparation. Membrane currents, electrical activity, and exocytosis were examined by patch-clamp technique measurements. Intracellular calcium concentration ([Ca(2+)](i)) was measured by microfluorimetry. Total internal reflection fluorescence microscope (TIRFM) was used for optical imaging of exocytosis and submembrane depolarization-evoked [Ca(2+)](i). The functional data were complemented by analyses of histology and gene transcription.


RESULTS
After 15 weeks, but not 3 weeks, mice on HFD exhibited hyperglycemia and hypoinsulinemia. Pancreatic islet content and beta-cell area increased 2- and 1.5-fold, respectively. These changes correlated with a 20-50% reduction of glucose-induced insulin secretion (normalized to insulin content). The latter effect was not associated with impaired electrical activity or [Ca(2+)](i) signaling. Single-cell capacitance and TIRFM measurements of exocytosis revealed a selective suppression (>70%) of exocytosis elicited by short (50 ms) depolarization, whereas the responses to longer depolarizations were (500 ms) less affected. The loss of rapid exocytosis correlated with dispersion of Ca(2+) entry in HFD beta-cells. No changes in gene transcription of key exocytotic protein were observed.


CONCLUSIONS
HFD results in reduced insulin secretion by causing the functional dissociation of voltage-gated Ca(2+) entry from exocytosis. These observations suggest a novel explanation to the well-established link between obesity and diabetes.
Oxford Centre for Diabetes, Endocrinology and Metabolism, University of Oxford, Churchill Hospital, Oxford, UK. stephan.collins@drl.ox.ac.uk
Stephan C
SC
Collins
Michael B
MB
Hoppa
Jonathan N
JN
Walker
Stefan
S
Amisten
Fernando
F
Abdulkader
Martin
M
Bengtsson
Jane
J
Fearnside
Reshma
R
Ramracheya
Ayo A
AA
Toye
Quan
Q
Zhang
Anne
A
Clark
Dominique
D
Gauguier
Patrik
P
Rorsman
D000818:Animals; D015220:Calcium Channels / Q000378:metabolism*; D003924:Diabetes Mellitus, Type 2 / Q000139:chemically induced* / Q000378:metabolism / Q000503:physiopathology*; D004041:Dietary Fats / Q000009:adverse effects*; D005089:Exocytosis / Q000502:physiology; D005434:Flow Cytometry; D018149:Glucose Intolerance / Q000378:metabolism; D007150:Immunohistochemistry; D007328:Insulin / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence; D022142:Secretory Vesicles / Q000378:metabolism*
D015220:Calcium Channels; D004041:Dietary Fats; D007328:Insulin