Background: During neurogenic inflammation, neuro–peptides (substance P, SP, calcitonin gene–related peptide, CGRP, and neurokinin A) are released from nerve endings, and these peptides initiate inflammatory reactions in the microcirculation. Platelets are one of the most important elements of the microcirculation. Methods: Our in vitro experiments were carried out to determine the effects of inflammatory neuropeptides (SP, and CGRP) on the arachidonate cascade of isolated rat platelets. Cells were labeled with 1–14C–arachidonic acid, then the eicosanoids were separated with overpressure thin–layer chromatography or high–performance liquid chromatography and were quantitatively determined with a liquid scintillation analyzer. Results: SP (10–9 and 10–8 mol/l) significantly increased the activity of the arachidonate cascade. The lipoxygenase pathway was significantly stimulated by SP (10–11, 10–9 and 10–8 mol/l), while the cyclooxygenase system was inhibited by 10–12 mol/l, and stimulated by 10–9 mol/l SP. The dose–response curve of TxA2 to SP exhibited a similar pattern to that detected for the cyclooxygenase pathway. Among the vasodilator cyclooxygenase metabolites, only the synthesis of PGE2 was significantly elevated by SP (10–9 mol/l ). CGRP either in low (10–12–10–11 mol/l) or in high concentrations (10–6 mol/l) activated the cyclooxygenase pathway, while it had no effect on the lipoxygenase pathway. CGRP (8–37), a specific CGRP1 receptor antagonist, inhibited the effects of CGRP. Conclusions: Our data suggest that the arachidonate metabolites of platelets may play a role in the process of neurogenic inflammation.

Hyperglycemia, hyperlipidemia, and free radicals result in platelet activation and atherogenesis. Kisspeptin (KP) is able to regulate metabolism, hemostasis, and the development of atherosclerosis. We examined whether platelet aggregation of streptozotocin-induced diabetic rats depends on the inducer type and if KP-13 and RF-9 (a kisspeptin receptor modifier) can influence platelet function. We measured the speed and the maximum of aggregation, along with the area under the curve. Serum glucose and calcium levels and urine formation of diabetic animals increased, while the body mass and platelet count decreased. Collagen was the most effective inducer of platelet aggregation. The aggregability of nondiabetic platelets was elevated in the presence of 5 × 10⁻⁸ mol/L KP-13. This effect was less expressed in diabetic animals. The effectivity of RF-9 was stronger than that of KP-13 in nondiabetic platelets, however it was ineffective in diabetic animals. RF-9 pre-treatment did not change the effects of 5 × 10⁻⁸ mol/L KP-13 in either animal group. The in vivo activation of diabetic platelets, which may be due to elevated serum calcium, induces thrombocytopenia and may lead to reduced in vitro aggregability. We could not demonstrate the antagonistic effect of RF-9 against KP-13 in isolated platelets.

Sigma-1 receptor (S1R) is detected in different cell types and can regulate intracellular signaling pathways. S1R plays a role in the pathomechanism of diseases and the regulation of neurotransmitters. Fluvoxamine can bind to S1R and reduce the serotonin uptake of neurons and platelets. We therefore hypothesized that platelets express S1R, which can modify platelet function. The expression of the SIGMAR1 gene in rat platelets was examined with a reverse transcription polymerase chain reaction and a quantitative polymerase chain reaction. The receptor was also visualized by immunostaining and confocal laser scanning microscopy. The effect of S1R agonist PRE-084 on the eicosanoid synthesis of isolated rat platelets and ADP- and AA-induced platelet aggregation was examined. S1R was detected in rat platelets both at gene and protein levels. Pretreatment with PRE-084 of resting platelets induced elevation of eicosanoid synthesis. The rate of elevation in thromboxane B₂ and prostaglandin D₂ synthesis was similar, but the production of prostaglandin E₂ was higher. The concentration-response curve showed a sigmoidal form. The most effective concentration of the agonist was 2 µM. PRE-084 increased the quantity of cyclooxygenase-1 as detected by ELISA. PRE-084 also elevated the ADP- and AA-induced platelet aggregation. S1R of platelets might regulate physiological or pathological functions.

To establish the possible influence of isatin (2,3-dioxo-indole) on the activity of platelets, the effects of isatin on platelet eicosanoid synthesis were studied in rats. Different doses (12.5–50 mg/kg) of isatin were injected intraperitoneally (i.p.) and the effects on the arachidonate cascade of isolated platelets were investigated. Cells were labeled with [¹⁴C]arachidonic acid, then the eicosanoids were separated with overpressure thin-layer chromatography and were quantitatively determined with a liquid scintillation analyzer. The lipoxygenase pathway was significantly inhibited by isatin (50 mg/kg) treatment and also the overall activity of the arachidonate cascade was diminished; however, the cyclooxygenase system was significantly stimulated. A 50-mg/kg i.p. dose of isatin significantly increased the production of vasoconstrictor cyclooxygenase metabolites. Among the vasodilator cyclooxygenase products, the synthesis of PGE2 and PGD2 were significantly decreased while that of 12-hydroxyheptadecatrienoic acid (HHT) increased upon isatin (50 mg/kg) administration. Our results provide further evidence on the peripheral actions of isatin and suggest that this endogenous indole may induce significant changes in the production of blood platelet arachidonic acid metabolites, which are important regulatory substances, thus isatin may potentially affect an even broader range of functions than was previously assumed.