Analysis of acid excretion in fish and its application to acidic environments.

The discovery in the late 1960s of ‘dead’ or dying lakes (Beamish and Harvey, 1972) nearly devoid of living animals and plants owing to the effects of acid precipitation spawned intense research into the effects of low pH environments on the physiology of freshwater fish. Over the next decade, an extensive body of literature documented the adverse effects of acid environments on acid–base status, ion levels and oxygen transport (Fromm, 1980). It became abundantly clear that acid exposure killed fish, but how acid killed fish, i.e. the underlying mechanisms and the proximate cause of mortality of acid-exposed fish, remained uncertain. Enter Chris Wood, a newly hired faculty member at McMaster University, Canada, and his collaborator, Gord McDonald. Wood and McDonald recognized the need for mechanistic studies of the ionoregulatory and acid–base disturbances of acid-exposed fish, and set out to generate the necessary data, adapting established experimental approaches to their needs. Their 1981 paper in The Journal of Experimental Biology (McDonald and Wood, 1981) showcased this new approach, yielding valuable insight into the mechanisms underlying acid–base disturbances in acid-exposed fish, as well as experimental methods for the measurement of acid–base fluxes that are still in use some 30years later. The rainbow trout used by McDonald and Wood were fitted with an arterial catheter, or cannula – to allow repetitive blood sampling without disturbing the fish – and then the blood samples were analyzed for acid–base status and ion concentrations. Crucially, the duo also recognized the value of analyzing the acid–base status and ion concentrations of the water in which the fish were held. By measuring changes in water ion concentrations and acid–base equivalents for fish held in a recirculating (i.e. closed) water system, McDonald and Wood were able to calculate net ion and acid–base fluxes, that is, values for the net exchange of ions and acid–base equivalents between the animal and its environment. In fish, both the gill and the kidney are key sites of ionic and acid–base regulation. Thus, McDonald and Wood fitted the fish with a bladder catheter for urine collection. Separate analyses of the collected urine and the water in which the fish were held allowed the relative contributions of the gill and kidney to net ion and acid–base fluxes to be distinguished. McDonald and Wood therefore were able to measure gains or losses of ions and acid across the gill and via the urine, and to assess the importance of these transfers to blood acid–base and ion status over time as the fish were exposed to acidic water (pH4). In so doing, they identified the gill as the site of acid uptake and demonstrated an important role for the kidney in excreting the acid load gained via the gill. Renal acid excretion was not, however, sufficient to compensate for branchial acid gain, resulting in a depression of blood pH. They also reported that exposure to acid water promoted a continuous loss of ions across the gill, accounting for the well-known lowering of plasma ion levels in acid-exposed fish, and they suggested that fluid shifts triggered by ion losses might ultimately cause circulatory collapse and death, a hypothesis supported by subsequent studies (e.g. Milligan and Wood, 1982).