Effect of platelet-activating factor (PAF) on human platelets

The effect of pure synthetic PAF (1-0-alkyl-2-acetyl-sn-glycero-3- phosphorylcholine) was studied in human platelets. PAF (0.2--2.0 micrograms/ml) produced a dose-dependent aggregation in human platelet- rich plasma (PRP) or platelet suspension obtained by gel-filtration (GFP). In addition, PAF (0.8 microgram/ml) induced secretion of 14C- serotonin (45% +/- 10%; mean +/- SD, n = 9) and platelet factor 4 (PF4) (12.89 +/- 3.81 micrograms/10(9) platelets; n = 9) in PRP. Similar results were obtained in GFP. Aggregation and release of 14C-serotonin and PF4 were inhibited by the metabolic inhibitors 2-deoxyglucose (16.7 mM) and antimycin-A (8.3 micrograms/ml), by the membrane-active drugs mepacrine (10 microM) and chlorpromazine (0.025 mM), by PGI2 (5.34 nM), which elevates intracellular c-AMP, by indomethacin (10 microM) or aspirin (100 microM). The ADP scavengers, creatine phosphate and creatine phosphokinase (CP/CPK), inhibited the second wave of aggregation but not secretion. These data suggest that the major effect of PAF on human platelets is mediated through the cyclo-oxygenase pathway and not through a third pathway.     

Red cell compatibility testing in baboon xenotransplantation

BACKGROUND: Although baboon ABO group and human anti-baboon heteroagglutinin (HA) titers have been considered in the selection of baboon donors for clinical hepatic xenotransplantation, the biologic role of these antibodies is not yet known. However, because of the potential importance of ABO hemagglutinins, a method for baboon ABO group determination is described, as are the titers of HA observed in both baboons and normal human donors. STUDY DESIGN AND METHODS: The ABO group of 62 baboons was determined by modified reverse typing. Baboon sera were heated and absorbed with human group O red cells. Reverse typing was then performed by standard techniques. HA titers at room temperature (RT) and in the antiglobulin test (AGT) were assessed in 10 baboons by using human red cells and in 33 normal donors by using baboon red cells. RESULTS: Ten (16%) baboons were group A, 29 (47%) were group B, 23 (37%) were group AB, and none were group O. In tests using human group O red cells, HA titers in 10 baboons ranged from 1 to 32 at RT and from negative to 64 in the AGT. All 33 normal human sera contained anti-baboon HA. Under a hemagglutination scoring system, group A persons had the lowest HA scores (17 +/− 15 at RT, 31 +/− 19 in the AGT), and group B persons had the highest HA scores (67 +/− 4 at RT, 85 +/− 9 in the AGT). CONCLUSION: Baboon ABO group can be easily determined by modified reverse serum typing. Both baboons and humans possess HAs of variable titer. Among humans, titers appear to be highest in group B individuals and lowest in group A. Additional studies are needed to determine the clinical significance of these antibodies.     

The association of Triatoma maculata (Ericsson 1848) with the gecko Thecadactylus rapicauda (Houttuyn 1782) (Reptilia: Squamata: Gekkonidae): a strategy of domiciliation of the Chagas disease peridomestic vector in Venezuela?

Objective

To investigate the bioecological relationship between Chagas disease peridomestic vectors and reptiles as source of feeding.

Methods

In a three-story building, triatomines were captured by direct search and electric vacuum cleaner search in and outside the building. Then, age structure of the captured Triatoma maculata (T. maculata) were identified and recorded. Reptiles living in sympatric with the triatomines were also searched.

Results

T. maculata were found living sympatric with geckos (Thecadactylus rapicauda) and they bit residents of the apartment building in study. A total of 1 448 individuals of T. maculata were captured within three days, of which 74.2% (1 074 eggs) were eggs, 21.5% were nymphs at different stages, and 4.3% were adults.

Conclusions

The association of T. maculata and T. rapicauda is an effective strategy of colonizing dwellings located in the vicinity of the habitat where both species are present; and therefore, could have implications of high importance in the intradomiciliary transmission of Chagas disease.     

Negative cooperativity in binding of muscarinic receptor agonists and GDP as a measure of agonist efficacy

BACKGROUND AND PURPOSE

Conventional determination of agonist efficacy at G-protein coupled receptors is measured by stimulation of guanosine-5′-γ−thiotriphosphate (GTPγS) binding. We analysed the role of guanosine diphosphate (GDP) in the process of activation of the M₂ muscarinic acetylcholine receptor and provide evidence that negative cooperativity between agonist and GDP binding is an alternative measure of agonist efficacy.

EXPERIMENTAL APPROACH

Filtration and scintillation proximity assays measured equilibrium binding as well as binding kinetics of [³⁵S]GTPγS and [³H]GDP to a mixture of G-proteins as well as individual classes of G-proteins upon binding of structurally different agonists to the M₂ muscarinic acetylcholine receptor.

KEY RESULTS

Agonists displayed biphasic competition curves with the antagonist [³H]-N-methylscopolamine. GTPγS (1 µM) changed the competition curves to monophasic with low affinity and 50 µM GDP produced a similar effect. Depletion of membrane-bound GDP increased the proportion of agonist high-affinity sites. Carbachol accelerated the dissociation of [³H]GDP from membranes. The inverse agonist N-methylscopolamine slowed GDP dissociation and GTPγS binding without changing affinity for GDP. Carbachol affected both GDP association with and dissociation from G_i/o G-proteins but only its dissociation from G_s/olf G-proteins.

CONCLUSIONS AND IMPLICATIONS

These findings suggest the existence of a low-affinity agonist-receptor conformation complexed with GDP-liganded G-protein. Also the negative cooperativity between GDP and agonist binding at the receptor/G-protein complex determines agonist efficacy. GDP binding reveals differences in action of agonists versus inverse agonists as well as differences in activation of G_i/o versus G_s/olf G-proteins that are not identified by conventional GTPγS binding.