Electrochemical patterns during Drosophila oogenesis: ion-transport mechanisms generate stage-specific gradients of pH and membrane potential in the follicle-cell epithelium

Abstract

Background: Alterations of bioelectrical properties of cells and tissues are known to function as wide-ranging signals during development, regeneration and wound-healing in several species. The Drosophila follicle-cell epithelium provides an appropriate model system for studying the potential role of electrochemical signals, like intracellular pH (pH_i) and membrane potential (V_mem), during development. Therefore, we analysed stage-specific gradients of pH_i and V_mem as well as their dependence on specific ion-transport mechanisms.

Results: Using fluorescent indicators, we found distinct alterations of pH_i- and V_mem-patterns during stages 8 to 12 of oogenesis. To determine the roles of relevant ion-transport mechanisms in regulating pH_i and V_mem and in establishing stage-specific antero-posterior and dorso-ventral gradients, we used inhibitors of Na⁺/H⁺-exchangers and Na⁺-channels (amiloride), V-ATPases (bafilomycin), ATP-sensitive K⁺-channels (glibenclamide), voltage-dependent L-type Ca²⁺-channels (verapamil), Cl^--channels (9-anthroic acid) and Na⁺/K⁺/2Cl^--cotransporters (furosemide). Either pH_i or V_mem or both parameters were affected by each tested inhibitor. While the inhibition of Na⁺/H⁺-exchangers (NHE) and amiloride-sensitive Na⁺-channels or of V-ATPases resulted in relative acidification, inhibiting the other ion-transport mechanisms led to relative alkalisation. The most prominent effects on pH_i were obtained by inhibiting Na⁺/K⁺/2Cl^--cotransporters or ATP-sensitive K⁺-channels. V_mem was most efficiently hyperpolarised by inhibiting voltage-dependent L-type Ca²⁺-channels or ATP-sensitive K⁺-channels, whereas the impact of the other ion-transport mechanisms was smaller. In case of very prominent effects of inhibitors on pH_i and/or V_mem, we also found strong influences on the antero-posterior and dorso-ventral pH_i- and/or V_mem-gradients. For example, inhibiting ATP-sensitive K⁺-channels strongly enhanced both pH_i-gradients (increasing alkalisation) and reduced both V_mem-gradients (increasing hyperpolarisation). Similarly, inhibiting Na⁺/K⁺/2Cl^--cotransporters strongly enhanced both pH_i-gradients and reduced the antero-posterior V_mem-gradient. To minor extents, both pH_i-gradients were enhanced and both V_mem-gradients were reduced by inhibiting voltage-dependent L-type Ca²⁺-channels, whereas only both pH_i-gradients were reduced (increasing acidification) by inhibiting V-ATPases or NHE and Na⁺-channels.

Conclusions: Our data show that in the Drosophila follicle-cell epithelium stage-specific pH_i- and V_mem-gradients develop which result from the activity of several ion-transport mechanisms. These gradients are supposed to represent important bioelectrical cues during oogenesis, e.g., by serving as electrochemical prepatterns in modifying cell polarity and cytoskeletal organisation.

Keywords: Bioelectricity; Cell polarity; Drosophila melanogaster; Epithelium; Follicle cell; Intracellular pH; Ion channel; Ion pump; Membrane potential; Pattern formation.