Platelet alpha-granules contribute to organ-specific pathologies in a mouse model of severe malaria

Cerebral malaria (CM) and malaria-associated acute lung injury/acute respiratory distress syndrome (MA-ALI/ARDS) are among the most severe complications of Plasmodium infection. While these disease manifestations are multifactorial, platelets have been described to play a role in the development of both syndromes in humans1,2 and mice3,4. Although the impact of platelets on malaria has been well-studied, questions remains with regard to their contribution to parasite control and immunopathogenesis. Studies have indicated that platelets can kill Plasmodium-infected red blood cells (iRBCs)5-8. However, there are contrasting reports that platelets do not exert any significant control over parasite growth but rather exacerbate malaria immunopathology3,9-12. In this study, we address the role of platelets in the development of severe malaria in three different mouse models of platelet dysfunction/depletion. We show a key role for platelets, and particularly platelet alpha granules (-granules), in mediating organ-specific pathologies during rodent Plasmodium infection

Heterologous Expression in Remodeled C. elegans: A Platform for Monoaminergic Agonist Identification and Anthelmintic Screening.

Monoamines, such as 5-HT and tyramine (TA), paralyze both free-living and parasitic nematodes when applied exogenously and serotonergic agonists have been used to clear Haemonchus contortus infections in vivo. Since nematode cell lines are not available and animal screening options are limited, we have developed a screening platform to identify monoamine receptor agonists. Key receptors were expressed heterologously in chimeric, genetically-engineered Caenorhabditis elegans, at sites likely to yield robust phenotypes upon agonist stimulation. This approach potentially preserves the unique pharmacologies of the receptors, while including nematode-specific accessory proteins and the nematode cuticle. Importantly, the sensitivity of monoamine-dependent paralysis could be increased dramatically by hypotonic incubation or the use of bus mutants with increased cuticular permeabilities. We have demonstrated that the monoamine-dependent inhibition of key interneurons, cholinergic motor neurons or body wall muscle inhibited locomotion and caused paralysis. Specifically, 5-HT paralyzed C. elegans 5-HT receptor null animals expressing either nematode, insect or human orthologues of a key Gαo-coupled 5-HT1-like receptor in the cholinergic motor neurons. Importantly, 8-OH-DPAT and PAPP, 5-HT receptor agonists, differentially paralyzed the transgenic animals, with 8-OH-DPAT paralyzing mutant animals expressing the human receptor at concentrations well below those affecting its C. elegans or insect orthologues. Similarly, 5-HT and TA paralyzed C. elegans 5-HT or TA receptor null animals, respectively, expressing either C. elegans or H. contortus 5-HT or TA-gated Cl- channels in either C. elegans cholinergic motor neurons or body wall muscles. Together, these data suggest that this heterologous, ectopic expression screening approach will be useful for the identification of agonists for key monoamine receptors from parasites and could have broad application for the identification of ligands for a host of potential anthelmintic targets

The 5-HT/SER-4-dependent inhibition of either the AIB interneurons or cholinergic motor neurons causes locomotory paralysis.

<p><b>A.</b> Confocal images of 5-HT <i>quint</i> expressing SER-4::GFP in the AIB interneurons (P<i>npr-9</i>)(A1) or cholinergic motor neurons (P<i>unc-17β</i>)(A2). GFP fluorescence (A2) or GFP fluorescence overlaid on DIC image (A1). The red stain in A2 is coelomocyte-specific RFP screening marker. <b>B.</b> Paralysis of wild type, mutant and transgenic <i>C</i>. <i>elegans</i> on hypotonic, non-NGM agar plates. Wild type, quadruple 5-HT receptor null animals expressing only SER-4 (SER-4 <i>quad</i>) or 5-HT <i>quint</i> expressing the <i>C</i>. <i>elegans</i> 5-HT₁-like receptor, SER-4, in either the cholinergic motor neurons (P<i>unc-17β</i>) or the two AIB interneurons (P<i>npr-9</i>) were examined for 5-HT (1 mM)-dependent paralysis as outlined in Methods. Data are presented as mean ± SE (n = 3).</p