An animal model to study class D L-type Ca 2+ channels

Insulin-secretion in pancreatic beta-cells is mainly controlled by Ca2+ influx through voltage-gated L-type Ca2+ channels Drugs that selectively modulated the function of L-type Ca2+ channels in pancreatic beta-cells are considered promising new therapeutic tools for the treatment of type I and II diabetics. Currently available L-type Ca2+ channel modulators, such as dihydropyridines (DHPs), cannot be exploited for this therapeutic goal, because of their preferential action in the cardiovascular system in vivo. Therefore the potential therapeutic effects of L- type channel modulation in pancreatic beta-cells can only be assessed and exploited by developing a new generation of L-type channel modulators that selectively target L-type channels in endocrine tissues and lack cardiovascular effects. Recent studies have provided a rational basis for the development of such drugs: L-type Ca2+ channels in cardiovascular muscle and pancreatic beta-cells are formed by different channel isoforms: in vascular smooth and cardiac muscle only so-called class C channels (containing a DHP-sensitive alpha-1C subunit) are expressed. In contrast, in pancreatic beta-cells the class D isoform (containing a DHP-sensitive alpha-1D subunit) is the predominant form. We therefore hypothesize that class D selective drugs could represent such a new generation of endocrine-selective drugs modulating Ca2+ influx and insulin-secretion in beta-cells without affecting the cardiovascular system. To test our hypothesis we propose an animal model that can be used to assess the therapeutic potential of class D channel modulation in vivo without the need to develop class D selective drugs. This will be accomplished by targeted disruption of DHP sensitivity of class C channels in mice. The rationale is to introduce one or several targeted mutations (in channel segments IIIS5 or IVS6) into the mouse alpha-1C-gene thereby reducing high DHP sensitivity for channel blockers (e.g. isradipine) and activators. (e.g. BAYK 8644). In homozygous mice class C channel complexes should display negligible DHP sensitivity and therefore DHP effects should be limited to class D Ca2+ channels Thus in these mice currently available DHPs should act as class D - selective modulators. As class D channels are also expressed to various extents in other endocrine cells and neurons this animal model will provide an important tool to study their contribution to L-type Ca2+ currents and their physiological role in these tissues. In vitro and in vivo analysis should allow to predict the therapeutic potential of selective class D channel modulators for the treatment of diabetes mellitus, the potential side effects of such drugs and may also reveal even other interesting therapeutic indications by modulation of Ca2+ dependent processes in other endocrine cells or in the central nervous system. The development of this mouse model will be complemented by an extensive analysis of the biochemical, pharmacological and modulatory properties of class D Ca2+ channels in neuronal and endocrine tissues and after expression in heterologous systems.