Active excretion of ammonia across the gills of the shore crab Carcinus maenas and its relation to osmoregulatory ion uptake

The mechanism of transbranchial excretion of total ammonia of brackish-water acclimated shore crabs, Carcinus maenas was examined using isolated, perfused gills. Applying physiological gradients of NH4Cl (100-200 mu mol.l(-1)) directed from the haemolymph space to the bath showed that the efflux of total ammonia consisted of two components. The saturable component (excretion of NH4+) greatly exceeded the linear component (diffusion of NH3). When an outwardly directed gradient (200 mu mol.l(-1)) was applied, total ammonia in the perfusate was reduced by more than 50% during a single passage of saline through the gill. Effluxes of ammonia along the gradient were sensitive to basolateral dinitrophenol, ouahain, and Cs+ and to apical amiloride. Acetazolamide (1 mmol.l(-1) basolateral) or Cl--free conditions had no substantial effects on ammonia flux, which was thus independent of both carbonic anhydrase mediated pH regulation and osmoregulatory NaCl uptake. When an inwardly directed gradient (200 mu mol l(-1)) was employed, influx rates were about 10-fold smaller and unaffected by basolateral ouabain (5 mmol.l(-1)) or dinitrophenol (0.5 mmol.l(-1)). Under symmetrical conditions (100 mu mol.l(-1) NH4Cl on both sides) ammonia was actively excreted against the gradient of total ammonia, which increased strongly during the experiment and against the gradient of the partial pressure of NH3. The active excretion rate was reduced to 7% of controls by basolateral dinitrophenol (0.5 mmol.l(-1)), to 44% by basolateral ouabain (5 mmol.l(-1)), to 46% by Na+-free conditions and to 42% by basolateral Cs+ (10 mmol.l(-1)), indicating basolateral membrane transport of NH4+ via the Na+/K+-ATPase and K+-channels and a second active, K+-ATPase and K+-channels and a second active, apically located, Na+ independent transport mechanism of NH4+. Anterior gills, which are less capable of active ion uptake than posterior gills, exhibited even increased rates of active excretion of ammonia. We conclude that, under physiological conditions, branchial excretion of ammonia is a directed process with a high degree of effectiveness. It even allows active extrusion against an inwardly directed gradient, if necessary.