Inhibitory effects of amitriptyline, sertraline and chlorpromazine on the thrombin-induced aggregation of platelets

Platelets resemble monoaminergic neurons in several respects, i.e the uptake of 5-HT and its inhibition, the subcellular storage and release of 5-HT, and the metabolism of aromatic amines brought about by monoamine oxidase. And the 5-HT content of rabbit platelets is well known to be about 40 times higher than that of human platelets. Therefore, this study was carried out to investigate the influences of amitriptyline (AMT) and sertraline (SRT) on the aggregation, contents of signaling second messengers, and protein phosphorylation of rabbit platelets in response to thrombin, 0.25 unit/ml, comparing with those of chlorpromazine (CPZ). Thrombin-induced aggregation was inhibited by SRT (IC50: 4.37 x 10^-5 M), CPZ (IC50: 5.76 x 10^-5 M), and AMT (IC50: 1.15 x 10^-4 M), respectively, and the aggregation by A23187 (1.0 μM) or PMA (320 nM) was also inhibited by SRT, CPZ, and AMT. AMT, SRT, and CPZ had little affects on basal contents of platelet TXB₂ and PGE₂, but all of them inhibited the thrombin-induced increase of TXB₂. Thrombin did not change the platelet contents of cAMP and cGMP. CPZ, AMT, and SRT produced the slight decrease of basal cAMP content, and their effects were not affected by thrombin-treatment. But SRT and AMT moderately increased the basal cGMP content, and the cGMP content of thrombin-stimulated platelets was gradually increased by the pretreatment with SRT, AMT, and CPZ. Particularly, the SRT-dependent increase of the cGMP content was notable. Platelet Ins(1,4,5)P₃ content was rapidly increased up to a plateau within 10 sec after thrombin-stimulation. AMT, SRT, and CPZ increased the basal Ins(1,4,5)P₃ content, and the thrombin-dependent increase was enhanced by pretreatment with CPZ and AMT, but was blunted by SRT. Platelet [Ca²⁺](i) was rapidly increased up to a peak level within 20 sec after thrombin-stimulation. The increase of [Ca²⁺]i was significantly inhibited by AMT, SRT, and CPZ. Thrombin- or PMA-induced phosphorylations of platelet 41 ~ 43 kDa and 20 kDa proteins were significantly inhibited by AMT, SRT, and CPZ. These results suggest that the antiplatelet activities of AMT and CPZ may be considerably attributed to the inhibition of protein kinase C activity, and the activity of SRT may be associated with the inhibitory effect on the thrombin-induced increase of Ins(1,4,5)P₃ and the increasing effect on the cGMP content of platelets. Therefore, it seems to be evident that AMT and SRT may produce their antidepressant activity, at least, partly through the inhibition of protein kinase C activity or the increase of resting Ins(1,4,5)P₃ content and in case of SRT, to a lesser extent, via the increase of cGMP in the brain.