Chapter 25 Insulin Secretion from β-Cells is Affected by Deletion of Nicotinamide Nucleotide Transhydrogenase
Shimomura K., Galvanovskis J., Goldsworthy M., Hugill A., Kaizak S., Lee A., Meadows N., Quwailid MM., Rydström J., Teboul L., Ashcroft F., Cox RD.
Nicotinamide nucleotide transhydrogenase (NNT) is an inner mitochondrial membrane transmembrane protein involved in regenerating NADPH, coupled with proton translocation across the inner membrane. We have shown that a defect in Nnt function in the mouse, and specifically within the β-cell, leads to a reduction in insulin secretion. This chapter describes methods for examining Nnt function in the mouse. This includes generating in vivo models with point mutations and expression of Nnt by transgenesis, and making in vitro models, by silencing of gene expression. In addition, techniques are described to measure insulin secretion, calcium and hydrogen peroxide concentrations, membrane potential, and NNT activity. These approaches and techniques can also be applied to other genes of interest. © 2009 Elsevier Inc. All rights reserved.