Protection by Natural Blackwater against Disturbances in Ion Fluxes Caused by Low pH Exposure in Freshwater Stingrays Endemic to the Rio Negro

Abstract

Stenohaline freshwater stingrays (Potamotrygon spp.) are endemic to the very dilute (Na(+), Cl(-), Ca2(+) <or=30 micromol L(-1)), often acidic blackwaters of the Rio Negro despite gill Na(+) and Cl(-) transport characteristics that appear unfavorable (high K(m), low J(max)). We evaluated the possible protective role of blackwater itself, which is rich in dissolved organic carbon (DOC), as well as the importance of Ca(2+) in allowing this tolerance of dilute, acidic conditions. Responses of stingrays in natural blackwater (DOC=8.4 mg L(-1)) were compared with those in a natural reference water with similar ion levels but low DOC (0.6 mg L(-1)). Comparing these two water types, we found that differences in Na(+) and Cl(-) unidirectional fluxes (JXin, JXout; measured with radiotracers) and net fluxes (JXnet), influx and outflux kinetic relationships, and net ammonia excretion (J(Amm)) were generally small at pH 6.3, though the balance points where Jin=Jout shifted from >300 micromol L(-1) in reference water (low DOC) to about 100 micromol L(-1) in blackwater (high DOC). In reference water, both JNain and JClin were inhibited >90%, both JNaout and JClout more than doubled, and J(Amm) did not change at pH 4.0. In blackwater, the inhibition of influxes was attenuated, the increases in outflux did not occur, and J(Amm) increased by 60% at pH 4.0. Addition of 100 micromol L(-1) Ca(2+) to reference water prevented the increases in JNaout and JClout and allowed J(Amm) to increase at pH 4.0, which demonstrates that the gills are sensitive to Ca(2+). However, addition of Ca(2+) to blackwater had no effect on the responses to pH 4.0. Addition of commercial humic acid to reference water did not duplicate the effects of natural Rio Negro blackwater at the same DOC level; instead, it greatly exacerbated the increases in JNaout and JClout at low pH and prevented any protective influence of added Ca(2+). Thus, blackwater DOC appears to be very different from commercial humic acid. Biogeochemical modeling indicated that blackwater DOC prevents Ca(2+) binding, but not H(+) binding, to the gills and that the protective effects of blackwater cannot be attributed to its higher buffer capacity or its elevated Al or Fe levels. Natural DOC may act directly at the gills at low pH to exert a protective effect and, when doing so, may override any protective action of Ca(2+) that might otherwise occur.

Growth of silver catfish ( Rhamdia quelen ) exposed to acidic pH at different humic acid levels

“…Humic substances may be stimulant and protector of paracellular junctions of the gill epithelium, thus preventing ion efflux in some fish from the natural acidic black waters of Rio Negro (GONZALEZ et al, 2002;2005;WOOD et al, 2003). This protective action could spare energy to be used in fish growth, explaining the greater development of green sword tail exposed to 180mg L -1 humic substance compared to control (0mg L -1 humic substance) (MEINELT et al, 2004).…”

“…The organic compounds of humic substances result from decomposition of dead plants and animals and contain acidic functional groups, besides being important regulators of biogeochemical processes, such as global nutrients and carbon cycle (WOOD et al, 2003;MATSUO & VAL, 2007). Black waters of the Amazon region are known for their physicochemical characteristics, relatively uncommon to average global values, with particular respect to the acidic pH parameters (it can reach 3.0 in some situations) and low ionic concentration.…”

“…II action on the gills, exerting a protective effect at low pH and, being able to replace any protective property that Ca 2+ could possess (WOOD et al, 2003). Fish kept in natural black waters, rich in humic substances, show a lower ion efflux after exposure to low pH levels than fish maintained in water without humic substance, suggesting that it reduce gill permeability, preventing ion loss by diffusion (GONZALES et al, 2002;2005;WOOD et al, 2003). Furthermore, humic substances facilitate the secondary repair of lesions induced by stress in fish (MEINELT et al, 2008).…”

“…A study with green swordtail (Xiphophorus helleri) has been the only to report some beneficial effect on fish growth for humic substances (MEINELT et al, 2004). Humic substances apparently have a protective effect on osmoregulation in some fish species exposed to acidic waters (GONZALES et al, 2002;2005;WOOD et al, 2003), but not in silver catfish (COSTA et al, 2015). Due to the relevant commercial importance of silver catfish in the south of Brazil, this study aimed to evaluate the effect of different concentrations of humic acid (HA) (a fraction of humic substance) on the species performance when exposed to different pHs.…”

Effect of humic acid on survival, ionoregulation and hematology of the silver catfish, Rhamdia quelen (Siluriformes: Heptapteridae), exposed to different pHs

“…The high levels of DOC account for the acidity of the aquatic environment. In order to thrive in acidic environments, organisms need a certain degree of specialization in their osmoregulatory organs (MATSUO & VAL 2007).Low pH (pH 4-5) induces ion loss (ZAIONS & BALDISSEROTTO 2000, 2003, BOLNER & BALDISSEROTTO 2007, MATSUO & VAL 2007, DUARTE et al 2013) and the interference with gill ionoregulatory mechanisms may also trigger hematological disturbances. Ionic dilution, potentiated by the plasma acidosis prompted by H + entry, affects body fluid distribution.…”