Adenovirus-mediated knockout of a conditional glucokinase gene in isolatedpancreatic islets reveals an essential role for proximal metabolic coupling events in glucose-stimulated insulin secretion
Dw. Piston et al., Adenovirus-mediated knockout of a conditional glucokinase gene in isolatedpancreatic islets reveals an essential role for proximal metabolic coupling events in glucose-stimulated insulin secretion, J BIOL CHEM, 274(2), 1999, pp. 1000-1004
The relationship between glucokinase (GK) and glucose-stimulated metabolism
, and the potential for metabolic coupling between beta cells, was examined
in isolated mouse islets by using a recombinant adenovirus that expresses
Cre recombinase (AdenoCre) to inactivate a conditional GK gene allele (gk(l
ox)). Analysis of AdenoCre-treated islets indicated that the gk(lox) allele
in similar to 30% of islet cells was converted to a nonexpressing variant
(gk(del)). This resulted in a heterogeneous population of beta cells where
GK was absent in some cells. Quantitative two-photon excitation imaging of
NAD(P)H autofluorescence was then used to measure glucose-stimulated metabo
lic responses of individual islet beta cells from gk(lox/lox) mice. In Aden
oCre-infected islets, approximately one-third of the beta cells showed mark
edly lower NAD(P)H responses. These cells also exhibited glucose dose respo
nses consistent with the loss of GK. Glucose dose responses of the low-resp
onding cells were not sigmoidal and reached a maximum at similar to 5 mM gl
ucose, In contrast, the normal response cells showed a sigmoidal response w
ith an KcatS0.5 of similar to 8 mM. These data provide direct evidence that
GK is essential for glucose-stimulated metabolic responses in beta cells w
ithin intact islets and that intercellular coupling within the islet plays
little or no role in glucose-stimulated metabolic responses.