Serotonin-stimulated calcium responses in human platelets: assay buffer dependency.
In Fura-2 labelled human platelets, the observed calcium response to serotonin stimulation is increased if the assay is conducted in a Krebs buffer compared with the HEPES buffer system usually used. The serotonin response in the Krebs buffer system was concentration dependent with an EC50 value of approximately 0.3 microM and was blocked by nanomolar concentrations of the serotonin2-receptor antagonist methiothepin. In a series of samples collected from 11 persons, a reasonable correlation was seen between the response to 1 microM serotonin and the serotonin2A-receptor density measured in corresponding membrane preparations. The calcium response to serotonin was reduced by treatment with hydrogen peroxide. It is concluded that the use of the Krebs buffer gives a more sensitive response of the Fura-2 loaded human platelets to serotonin stimulation than does the use of HEPES buffer and that the increased sensitivity is achieved without affecting the pharmacological properties of the response. For the response to thrombin, the response intensity is affected by the pH of the buffer used but is not sensitive to the buffer composition.
Fowler
C J
CJ
Department of Pharmacology, Umeå University, Sweden. cf@pharm.umu.se
Tiger
G
G
D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism*; D002021:Buffers; D020013:Calcium Signaling / Q000187:drug effects*; D005456:Fluorescent Dyes; D016257:Fura-2; D006531:HEPES; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D066298:In Vitro Techniques; D007552:Isotonic Solutions; D008719:Methiothepin / Q000494:pharmacology; D044402:Receptor, Serotonin, 5-HT2A; D011985:Receptors, Serotonin / Q000378:metabolism; D012701:Serotonin / Q000378:metabolism / Q000494:pharmacology*; D012702:Serotonin Antagonists / Q000494:pharmacology
D002021:Buffers; D005456:Fluorescent Dyes; D007552:Isotonic Solutions; C051865:Krebs-Ringer solution; D044402:Receptor, Serotonin, 5-HT2A; D011985:Receptors, Serotonin; D012702:Serotonin Antagonists; D012701:Serotonin; D008719:Methiothepin; D006861:Hydrogen Peroxide; D006531:HEPES; D016257:Fura-2