A medium termed 4suc:6KCl (containing 83mM sucrose, CC-5013 105mM K+, 6.8mM Na+, 66mM Cl?, and nutrients) supported parasite growth at rates matching those in standard RPMI 1640 medium. Because na?ve cultures expanded at normal rates immediately upon transfer to this medium, the parasite does not appear to require adaptation prior to expansion in this nonphysiological environment. We measured erythrocyte cation concentrations in trophozoite-stage infected cells after cultivation in 4suc:6KCl and found that host cation remodeling was fully prevented, consistent with passive channel-mediated movements of Na+ and K+ under physiological conditions. Because parasite growth was not measurably affected, these findings revealed that host cation remodeling is an unnecessary byproduct of PSAC activity.

This channel appears instead to be critical for nutrient acquisition. Use of nonphysiological media for in vitro cultivation has provided a number of other insights into parasite biology. For example, reducing Na+ below the above EC50 value produced trophozoites with engorged digestive vacuoles, suggesting parasite regulation of this ion within its compartments and possible new targets for intervention. Extracellular K+ also appears to be needed, albeit at low levels; this finding is surprising because one would have thought the sizeable erythrocyte K+ stores could adequately fulfill parasite demand. At the other end of the spectrum, parasite tolerance of K+ concentrations up to 148mM provides strong evidence against an essential role for K+ in merozoite maturation [31].

In another study using nonphysiological media, we found that parasites also have a broad tolerance to changes in extracellular pH [32], a desirable trait in light of the metabolic acidosis that often accompanies severe malaria [33]. Finally, through the use of sucrose as an osmoticant to replace salts in the media, we found that merozoite egress and invasion depend on a defined range of ionic strength values [30], suggesting electrostatic interactions between parasite macromolecules that may play critical roles in merozoite egress and invasion.4. ConclusionsIn vitro cultivation of P. falciparum, developed nearly 40 years ago, has enabled fundamental advances in both basic and clinical malaria research.

Over that time, a single RPMI 1640-based medium has achieved near universal use, leading to a number of assumptions about in vivo parasite behavior, drug action, prioritization Batimastat of parasite targets for future therapies, and parasite biology. Recent studies have used modifications to this standard medium to gain new insights into these and other questions. Additional manipulations of culture conditions are needed to identify new targets for therapeutic intervention and uncover conditions that permit cultivation of refractory plasmodial species.