Diversity and novel pharmacological properties of Ca2+ channels in Drosophila brain membranes.

Abstract

Binding studies as well as affinity labelling and immunoblot techniques were used to identify and characterize the receptors for Ca2+ channel blockers in Drosophila brain membranes. Despite structural analogies with mammalian receptors, Drosophila binding sites for phenylalkylamines and 1,4‐dihydropyridines, unlike those described in skeletal and cardiac muscle, were found to be located on separate Ca2+ channels. Single‐channel bilayer recordings from reconstituted membranes revealed the presence of eight distinct cobalt‐sensitive Ba2+‐conducting channels in Drosophila brain membrane preparations. In good agreement with binding studies, the most frequently observed Ca2+ channel type (Ba2+ conductance of 13 pS) was extremely sensitive to phenylalkylamines but not affected by micromolar concentrations of 1,4‐dihydropyridines. Distinct 1,4‐dihydropyridine‐sensitive and phenylalkylamine‐insensitive channels were also identified. They had unitary Ba2+ conductances of 21 and 31 pS. A detailed analysis of drug action showed that both 1,4‐dihydropyridines and phenylalkylamines first increased channel open state probability before fully blocking channel activity. Other types of channels have been identified with unitary Ba2+ conductances of 9, 41, 53, 64 and 81 pS. They were insensitive to the previously described organic Ca2+ channel blockers. The Drosophila system seems to be a unique model to analyse the properties of several different types of Ca2+ channels and particularly those of channel types that are uniquely blocked by phenylalkylamines or uniquely blocked by 1,4‐dihydropyridines.

Very high affinity interaction of DPI 201‐106 and BDF 8784 enantiomers with the phenylalkylamine‐sensitive Ca2+‐channel in Drosophila head membranes

“…Drosophila melanogaster head membranes contain at least eight distinct voltage-regulated Ca2+-channels after reconstitution in phospholipid bilayers (Pelzer et al, 1989). There is a Ca2" channel, permeable to Ba2" (13pS conductance) which is extremely sensitive to phenylalkylamines (transient stimulation and subsequent block by nm concentration of gallopamil or (-)-desmethoxyverapamil) but is completely insensitive to 1,4 dihydropyridines.…”

“…High-affinity phenylalkylamine binding sites are found in Drosophila melanogaster head membranes Pelzer et al, 1989;Greenberg et al, 1989). These sites are not coupled (as are mammalian L-type Ca2 +-channel phenylalkylamine receptors) to 1,4 dihydropyridine sites Greenberg et al, 1989) and can be specifically photolabelled by the arylazide phenylalkylamine [N-methyl-3H]-LU 49888 ((-)-5-[(3-azido-phenethyl)-[N-methyl-3H]methylamino-2-(3,4,5-trimethoxy-phenyl)-2 -isopropylvalero -nitrile)].…”

“…[N-methyl-3H]-LU 49888 photolabels a 135 K polypeptide in Drosophila head membranes Greenberg et al, 1989;Pelzer et al, 1989) with a sedimentation coefficient of 12 S upon sucrose gradient centrifugation after solubilization with digitonin (Greenberg et al, 1989). The Drosophila melanogaster phenylalkylamine-sensitive Ca2 +-channel seems to be unique, as Ca2+-antagonists from other chemical classes (benzothiazepines e.g.…”

Development of depolarization‐induced calcium transients in insect glial cells is dependent on the presence of afferent axons

“…In vertebrates, verapamil-sensitive L-type calcium channels are also sensitive to dihydropyridines such as nifedipine (Hille, 1992;Hockerman et al, 1997). In invertebrates, verapamil-sensitive calcium channels lack this sensitivity to dihydropyridines, while dihydropyridine-sensitive channels are insensitive to verapamil (Pelzer et al, 1989;Gielow et al, 1995;Takeuchi et al, 1996;Wicher and Penzlin, 1997;Morales et al, 1999;Beck et al, 2001). With only limited information about the molecular structure of invertebrate (mainly Drosophila) calcium channels, we are not yet able to classify the calcium channels by their pharmacologic properties.…”

Insect calcium channels

“…In arthropods different voltagegated calcium channels play a key role for muscle contraction: presynaptic channels control the release of excitatory transmitter (mainly glutamate) [lO,l l] at the neuromuscular junction, whereas calcium channels in the plasma membrane of muscle cells mediate the influx of calcium ions for contraction and participate in the propagation of the action potential [12]. The structural characterization of invertebrate calcium channels is rudimentary [13][14][15] and none of their subunits have yet been cloned.Here we report for the first time the cloning of an invertebrate calcium channel a,-subunit from Musea domestica larvae (designated Mdla,) that is preferentially expressed in body wall muscle. The evolutionary relationship of Mdla, compared to the so far cloned vertebrate calcium channels has been studied by calculating a phylogenetic tree.…”