Ammonia uptake and its effects on ionoregulation in the freshwater crayfish Pacifastacus leniusculus (Dana)
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abstract
Exposure of adult crayfish Pacifastacus leniusculus to Artificial Freshwater (AFW) media containing 1.5 m and 0.15 mmol x l(-1) total ammonia [Tamm; 0.1 x acute lethal concentration (24 h LC50) and 0.01 x 24 h LC50] and adjusted to pH 6.5, pH 8.2 and pH 10.5 resulted in significant increases in haemolymph ammonia over a 24-h period. Ammonia accumulated most rapidly at pH 10.5. These media were chosen to expose animals to a range of different un-ionised ammonia (UIA) [NH3] and ionised ammonia [NH4+] concentrations. From comparisons of measured transepithelial potential differences (PDte) with calculated Nernst potentials (PDNH4+) for the known haemolymph-to-medium gradients of [NH4+], it was deduced that, in pH 8.2 and pH 6.5 AFW, NH4+ was not in thermodynamic equilibrium across the integument (presumably gill epithelium). In pH 10.5 AFW with 1.5 mmol x l(-1) Tamm (predominantly NH3), the accumulation of ammonia in the haemolymph was in the NH4+ form due to haemolymph pH regulation by the crayfish in this alkaline external medium. Measured net fluxes of ammonia (Jamm(net)) were inwardly directed and maximal when [NH3] was the main component externally, but were also significant at pH 8.2 with high [NH4+] ([NH4+]:[NH3] approximately 20:1). Haemolymph Na+ depletion was significant and, over the 24-h exposure period, most rapid in high [NH3] medium but [Cl-] was unaffected. However, paradoxically, sodium uptake (measured JNa(in) on immediate transfer to high Tamm medium) was not significantly inhibited when [NH3] was the predominant ammonia species. In 1.5 mmol x l(-1) Tamm (mainly [NH4+), VNa(in) (the active component of JNa(in)) was significantly inhibited, particularly at low external [Na+]. This inhibition could not be demonstrated as one of competition at an Na+/NH4+ apical gill exchange site. The resultant net efflux of sodium from the animal showed that the ability of the animals to balance sodium losses at low external [Na+] was severely affected. Longer exposure to pH 10.5 AFW with high [NH3] (12 h) resulted in significantly increased JNa(out), while not significantly affecting JNa(in). Analysis of urinary Na+ losses showed that, while urinary flow rate and water reabsorption was most likely unaffected by ammonia exposure, final urine [Na+] was significantly elevated. The resulting urinary Na+ loss accounted for 63% of the increased JNa(out) in high [NH3] medium.
