PLATELET EXTERNAL SURFACE MEMBRANE IS OSMOTICALLY DOUBLED IRRESPECTIVE OF SIZE OR SPECIES (HUMAN/BOVINE): DYNAMICS AND MEMBRANE SOURCES

Abstract

Osmotic swelling can double the external plasma membrane surface area of human platelets independently of size, proposed to recruit the open surface-connected canalicular system ((SCCS) (Thrombos. Res. Suppl. VI: 119, 1986). As bovine (B) platelets have been reported to lack SCCS, we compared osmotic swelling for B and human (H) cells. Addition of water to piatelet-rich-plasma (10-90% v/v) caused sequential shape change and osmotic spherocyte (OS) formation, analyzed for size and surface area changes from time-dependent phase-contrast videomicroscopic images. Selected samples were fixed and stained with tannic acid prior to osmic acid fixation for visualization of open SCCS by transmission electron microscopy. B platelets required 3-4x less water dilution of PRP than H platelets, with significant OS forming at 20% water addition. Continued water dilution converted 50% of platelets to OS, with maximally stable swelling and no significant lysis for bovine OS up to 60% dilution. Electron micrographs of unactivated discocytes (D) and of optimally-swollen OS showed open SCCS in human D not detectable in any of the swollen platelets, though granules, mitochondria and a small number of vesicles and vacuoles persisted; no evidence for any open SCCS was found for bovine D or OS, though the OS otherwise appeared similar to H-0S. Geometric measurements of D and nonlysed OS showed a stable, maximal 2.1±0.1 fold increase in external plasma membrane surface area with osmotic swelling, identical for different-sized H platelets (mean volume = 2.8-6.8 f1) or for B platelets (3.6 f1 ). B platelets show equal or greater sensitivity for ADP-induced activation as H platelets, with 2-fold slower maximal rates of recruitment in early aggregation. As osmotic swelling appears to primarily externalize SCCS in H platelets, the identical relative amounts of internal membrane externalized for B platelets is hypothesized to arise from an osmotically more labile, “closed”, and structurally simpler SCCS or from a distinct membrane source tnan in H platelets.