Stimulation of protein phosphorylation in intact rat brain synaptosomes by a pyrethroid insecticide, deltamethrin

Abstract

Effects of deltamethrin, a powerful synthetic pyrethroid, on the protein phosphorylation activity in the intact rat brain synaptosome during the time course of depolarization-induced changes were studied. For this purpose, depolarization was induced either by veratridine or high concentration of K +. The level of phosphorylation on various synaptic proteins was found to quickly rise for 15 to 30 sec and return to the resting level in about 3 min. Deltamethrin, when given to the synaptosomes 10 min prior to depolarization, caused stimulation on this depolarization-induced protein phosphorylation activity at > 10 −12 M concentration. To find the cause for such an action of deltamethrin, the effects of ion-channel blockers and calmodulin inhibitors on the same phosphorylation process were studied. The inhibitory effect of a Ca 2+-channel blocker, verapamil, and a Na +-channel blocker, tetrodotoxin (TTX), on the stimulatory action of deltamethrin was first established. However, in neither case were their actions complete. The former reduced the stimulatory effect of deltamethrin on synapsin I but not on calcium- and calmodulin-dependent protein kinase II (CaM-Kinase II), B50, or 38-kDa proteins. The latter could not inhibit the stimulatory action of deltamethrin on synapsin I. Even when both blocking agents were present, the stimulatory action of deltamethrin was apparent. The stimulatory effect of deltamethrin on the phosphorylation rate of phosphoproteins, synapsin I, calcium-calmodulin-dependent protein kinase II and B50 was less when the entire external CaCl 2 was replaced by BaCl 2 in the prelabeling medium. Again, even under these conditions, the stimulatory effect of deltamethrin was evident on all proteins examined. In addition, the effect of calmodulin-inhibitor trifluoroperazine (TFP) was tested in the same manner. This inhibitor was found to reduce the phosphorylation on all of the tested synaptosome-proteins except protein 38 kDa, where there was an increase in phosphorylation in the presence of deltamethrin, especially, at 30, 60, and 180 sec of depolarization. These results indicate that (a) deltamethrin's action is not limited to calmodulin or the sodium or the calcium channel, (b) it must stimulate the release of Ca 2+ from the intracellular storage sites(s), and (c) such a stimulatory action of deltamethrin may be recognized only when the synaptosomes are depolarized regardless of the method of depolarization.