Addition of sialidase or p38 MAPK inhibitors does not ameliorate decrements in platelet in vitro storage properties caused by 4 °C storage.

BACKGROUND AND OBJECTIVES: Bacterial proliferation is inhibited in platelets (PLTs) stored at refrigerated temperatures, but also dramatically decreases PLT in vivo survival. Recent studies have demonstrated that cold temperature (CT) stored PLTs secrete sialidases upon re-warming, removing sialic acid from the PLT surface, which may be responsible for clustering of GPIbα and PLT clearance from circulation. In this study, the influence of a sialidase inhibitor or a p38 MAP kinase inhibitor was evaluated in units stored at 4 °C. MATERIALS AND METHODS: After collection of a single Trima apheresis unit (n = 12), PLTs were aliquoted into four 60-ml CLX storage bags. One bag was stored at 20-24 °C (RT) with continuous agitation; a second bag was stored at 4 °C without agitation; a third bag was held at 4 °C without agitation with sialidase inhibitor, a fourth bag was incubated at 4 °C with a p38 MAPK inhibitor without agitation. RESULTS: Beginning from Day 1, all in vitro PLT parameters were adversely affected by CT compared to those of RT. Similar in vitro storage properties were observed in CT PLT in the presence or absence of sialidase or p38 MAPK inhibitors. P38 MAPK phosphorylation inhibition was not observed at CT. Decrease of sialidase activity was observed for 2 days in PLTs stored in additive solution but not in plasma. CONCLUSION: Addition of either sialidase or p38 MAPK inhibitors do not improve any in vitro parameters of PLTs stored at 4 °C in 100% plasma.

Evaluation of in vitro storage properties of apheresis platelets suspended in a bicarbonate-containing additive solution with low levels of plasma and stored with a 24-hour interruption of agitation.

BACKGROUND AND OBJECTIVES: PLT additive solutions (PAS) are useful for reducing the frequency and/or severity of plasma-associated transfusion reactions. A new PAS solution, PAS-5, containing 5% plasma, maintains in vitro PLT properties during 7-day storage. Periods with interruption of agitation (IA) ≤24 h routinely occur during PLT shipment and do not usually compromise platelet quality. The aim of the study was to evaluate the properties of PLTs stored for 7 days in 95% PAS-5/5% plasma subjected to a 24-h IA. MATERIALS AND METHODS: Double apheresis Amicus units (n = 12) were collected using a manual PAS-5 addition to hyperconcentrated PLTs. PLT units were equally divided in two containers. Control and test PLTs were stored with continuous agitation at 20-24°C except for 24-h IA period for test units between days 2-3. RESULTS: During storage, levels of glucose, lactate, mitochondrial membrane potential and aggregation significantly differed in test units compared to those of control. The pH levels of test PLTs were less than those of control units with 7/12 test units having pHs <6·2 on Day 7 compared to 1/12 control units. Morphology score, GP1bα expression, ESC values, superoxide production were also less, and activation was greater in test PLTs than those of control. All other parameters were similar between test and control units. CONCLUSION: PLTs stored in PAS-5 solution containing 5% plasma with a 24-h IA results in marked decrements in many in vitro PLT quality parameters during 7-day storage.

Modulation of amphetamine-stimulated (transporter mediated) dopamine release in vitro by sigma2 receptor agonists and antagonists.

Some sigma receptor ligands have been shown to bind with low affinity to the dopamine transporter and to inhibit [3H]dopamine uptake. It has not previously been shown whether any of these compounds influence release of dopamine via facilitated exchange diffusion. To further examine the nature of the interaction between sigma receptor ligands and the dopamine transporter, the effects of sigma receptor ligands on amphetamine-stimulated [3H]dopamine release were examined in slices prepared from rat caudate putamen. In the absence of exogenous Ca2+, both (+)-pentazocine and (-)-pentazocine potentiated amphetamine-stimulated [3H]dopamine release at concentrations consistent with their affinities for sigma2 receptors. In contrast, BD737 (1S.2R-(-)-cis-N-¿2-(3,4-dichlorophenyl)ethyl¿-N-methyl-2-(1-pyrrolidiny l)cyclohexylamine), a sigma1 receptor agonist, had no effect on amphetamine-stimulated release. Neither isomer of pentazocine alone had any effect on basal [3H]dopamine release under these conditions. Three antagonists at sigma receptors, one of which is non-selective for subtypes, and two of which are sigma2-selective, all blocked the enhancement of stimulated release produced by (+)-pentazocine. Enhancement of stimulated release by (-)-pentazocine was similarly blocked by sigma2 receptor antagonists. Our data support the contention that it is possible to regulate transporter-mediated events with drugs that act at a subpopulation of sigma receptors pharmacologically identified as the sigma2 subtype.

Functional and binding properties of sigma receptors in rat cerebellum.

Autoradiographic studies have shown that sigma receptors are enriched in the locus coeruleus, the origin of noradrenergic projections to the cerebellum, as well as in the Purkinje, molecular, and granular layers and the interpositus cerebellar nucleus of the cerebellum itself. In contrast, the cerebellum is relatively poor in phencyclidine (PCP) binding sites, which have been historically confused with sigma sites. The high ratio of sigma to PCP receptors in cerebellum is advantageous for discriminating sigma-mediated physiological effects. sigma agonists and antagonists have been shown to regulate N-methyl-D-aspartate (NMDA)-stimulated norepinephrine release in hippocampus, which is innervated by locus coeruleus projections. We now report that sigma drugs also regulate norepinephrine release from cerebellum. In contrast to findings in the hippocampus, where regulation is via sigma 1 and sigma 2 receptors, sigma-mediated regulation in cerebellum seems to be primarily via sigma 1 receptors. In radioligand binding studies, we find that sigma receptors primarily of the sigma 1 type are present in the cerebellum. We further report that binding to sigma receptors in cerebellum is not affected by the addition of NMDA or glycine or by the presence of NMDA antagonists, suggesting that sigma receptors are not located within the NMDA-operated cation channel in this brain region.