Regulating your breathing can be tricky if you're a tambaqui swimming in the Amazon basin, where water CO2 levels change dramatically with the seasons. Wondering how the fish track these changes, Katie Gilmour and Steve Perry at the University of Ottawa decided to take a closer look at the location of tambaqui's CO2 receptors(p. 1095).
Gilmour and Perry travelled to Tadeu Rantin's lab in Brazil, where tambaqui experts Bill Milsom and Steve Reid joined the team. Collecting some tambaqui from a local fish farm, they set out to discover the orientation of tambaqui's CO2 receptors. The team reasoned that if tambaqui have internally oriented receptors, the fish should detect rising blood CO2 levels,but if they have externally oriented receptors, tambaqui should respond to CO2 levels in the surrounding water instead.
First, the team needed to prepare the fish so that they could measure tambaqui's cardiorespiratory responses to changing blood and water CO2 levels. They created an external blood flow loop, pulling blood from the artery that runs along the fish's back and returning it to the body,and inserted electrodes into the shunted bloodstream to take continuous blood gas measurements. But tambaqui have bony projections associated with their vertebral column, tucked away under the back muscles, and Gilmour remembers that it took `blood, sweat and tears to work cannula between these bones to set up the blood shunt'. Finally, they were ready to see how the fish handle fluctuating CO2 levels.
They began by testing tambaqui's responses to internal CO2changes. The team injected tambaqui with acetazolamide, which causes CO2 levels to build up in the bloodstream. If tambaqui have internal receptors that detect blood CO2 levels, the team expected to see ventilatory and cardiovascular responses in the acetazolamide-treated fish. But the treated fish didn't respond to their elevated blood CO2 levels at all; they didn't seem to have internal receptors.
To find out if the fish detect high water CO2 levels with external receptors instead, the team boosted water CO2 levels in the tanks of the acetazolamide-treated fish and then rapidly dropped the CO2 level back to normal again. If the fish have external receptors, the tambaqui's ventilatory and cardiovascular responses should track changes in water CO2 levels. Sure enough, as the team increased water CO2, they saw that the fish began hyperventilating and decreased their heart rate. And as water CO2 plummeted back to normal levels, they saw that the tambaqui's ventilation and heart rate returned to normal very quickly, despite the fact that these fish still had high levels of CO2 in their blood. Clearly, the fish were tracking changes in CO2 levels in the water but not in their blood. `We were amazed to see how beautifully ventilation and heart rate tracked water CO2 levels as they dropped, but didn't track blood CO2levels at all,' Gilmour recalls.
As a final demonstration of the orientation of tambaqui's receptors, the team injected CO2-enriched saline into the fish. They saw no response, bolstering their conclusion that tambaqui do not have internal receptors. But when they injected CO2-enriched water into the tambaqui's mouth, where it joined the flow of ventilatory water over the gills, they were pleased to see cardiorespiratory responses in the fish,confirming that tambaqui have external receptors located on their gills. So,when CO2 levels escalate in their Amazonian home, tambaqui's external receptors kick in straight away to help the fish cope with the respiratory challenge.