Regulation of extracellular ATP in human erythrocytes infected with Plasmodium falciparum

PLoS One. 2014 May 23;9(5):e96216. doi: 10.1371/journal.pone.0096216. eCollection 2014.

Abstract

In human erythrocytes (h-RBCs) various stimuli induce increases in [cAMP] that trigger ATP release. The resulting pattern of extracellular ATP accumulation (ATPe kinetics) depends on both ATP release and ATPe degradation by ectoATPase activity. In this study we evaluated ATPe kinetics from primary cultures of h-RBCs infected with P. falciparum at various stages of infection (ring, trophozoite and schizont stages). A "3V" mixture containing isoproterenol (β-adrenergic agonist), forskolin (adenylate kinase activator) and papaverine (phosphodiesterase inhibitor) was used to induce cAMP-dependent ATP release. ATPe kinetics of r-RBCs (ring-infected RBCs), t-RBCs (trophozoite-infected RBCs) and s-RBCs (schizont-infected RBCs) showed [ATPe] to peak acutely to a maximum value followed by a slower time dependent decrease. In all intraerythrocytic stages, values of ΔATP1 (difference between [ATPe] measured 1 min post-stimulus and basal [ATPe]) increased nonlinearly with parasitemia (from 2 to 12.5%). Under 3V exposure, t-RBCs at parasitemia 94% (t94-RBCs) showed 3.8-fold higher ΔATP1 values than in h-RBCs, indicative of upregulated ATP release. Pre-exposure to either 100 µM carbenoxolone, 100 nM mefloquine or 100 µM NPPB reduced ΔATP1 to 83-87% for h-RBCs and 63-74% for t94-RBCs. EctoATPase activity, assayed at both low nM concentrations (300-900 nM) and 500 µM exogenous ATPe concentrations increased approx. 400-fold in t94-RBCs, as compared to h-RBCs, while intracellular ATP concentrations of t94-RBCs were 65% that of h-RBCs. In t94-RBCs, production of nitric oxide (NO) was approx. 7-fold higher than in h-RBCs, and was partially inhibited by L-NAME pre-treatment. In media with L-NAME, ΔATP1 values were 2.7-times higher in h-RBCs and 4.2-times higher in t94-RBCs, than without L-NAME. Results suggest that P. falciparum infection of h-RBCs strongly activates ATP release via Pannexin 1 in these cells. Several processes partially counteracted ATPe accumulation: an upregulated ATPe degradation, an enhanced NO production, and a decreased intracellular ATP concentration.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphatases / metabolism
  • Adenosine Triphosphate / metabolism*
  • Biological Transport
  • Erythrocytes / cytology*
  • Erythrocytes / parasitology*
  • Extracellular Space / metabolism*
  • Homeostasis
  • Humans
  • Kinetics
  • Plasmodium falciparum / physiology*
  • Trophozoites / physiology

Substances

  • Adenosine Triphosphate
  • Adenosine Triphosphatases
  • ectoATPase

Grants and funding

This work was supported by the following agencies and organisations: CAPES-MINCYT (197/11), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET) Grant PIP 639, Secretaría de Ciencia y Técnica, Universidad de Buenos Aires Grant 20020100100090, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ), Coordenação de Aperfeiçoamento de Pessoal do Ensino Superior (CAPES), Agencia Nacional de Promoción Científica y Tecnológica Grant 0151 of Argentina and Instituto Nacional para Pesquisa Translacional em Saúde e Ambiente na Região Amazônica, Conselho Nacional de Desenvolvimento Científico e Tecnológico (INCT-INPeTAm/CNPq/MCT) of Brasil. CLA, SVV, PJS are career researchers from CONICET. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.