title: The relationship between NMDA receptor function and the high ammonia tolerance of anoxia-tolerant goldfish.
abstract: Acute ammonia toxicity in vertebrates is thought to be characterized by a cascade of deleterious events resembling those associated with anoxic/ischemic injury in the central nervous system. A key event is the over-stimulation of neuronal N-methyl-D-aspartate (NMDA) receptors, which leads to excitotoxic cell death. The similarity between the responses to acute ammonia toxicity and anoxia suggests that anoxia-tolerant animals such as the goldfish (Carassius auratus Linnaeus) may also be ammonia tolerant. To test this hypothesis, the responses of goldfish were compared with those of the anoxia-sensitive rainbow trout (Oncorhynchus mykiss Walbaum) during exposure to high external ammonia (HEA). Acute toxicity tests revealed that goldfish are ammonia tolerant, with 96 h median lethal concentration (LC(50)) values of 199 μmol l(-1) and 4132 μmol l(-1) for NH(3) and total ammonia ([T(Amm)]=[NH(3)]+[NH(4)(+)]), respectively. These values were ~5-6 times greater than corresponding NH(3) and T(Amm) LC(50) values measured in rainbow trout. Further, the goldfish readily coped with chronic exposure to NH(4)Cl (3-5 mmol l(-1)) for 5 days, despite 6-fold increases in plasma [T] to ~1300 μmol l(-1) and 3-fold increases in brain [T(Amm)] to 6700 μmol l(-1). Muscle [T(Amm)] increased by almost 8-fold from ~900 μmol kg(-1) wet mass (WM) to greater than 7000 μmol kg(-1) WM by 48 h, and stabilized. Although urea excretion rates (J(Urea)) increased by 2-3-fold during HEA, the increases were insufficient to offset the inhibition of ammonia excretion that occurred, and increases in urea were not observed in the brain or muscle. There was a marked increase in brain glutamine concentration at HEA, from ~3000 μmol kg(-1) WM to 15,000 μmol kg(-1) WM after 48 h, which is consistent with the hypothesis that glutamine production is associated with ammonia detoxification. Injection of the NMDA receptor antagonists MK801 (0.5-8 mg kg(-1)) or ethanol (1-8 mg kg(-1)) increased trout survival time by 1.5-2.0-fold during exposure to 2 mmol l(-1) ammonia, suggesting that excitotoxic cell death contributes to ammonia toxicity in this species. In contrast, similar doses of MK801 or ethanol had no effect on ammonia-challenged (8-9.5 mmol l(-1) T(Amm)) goldfish survival times, suggesting that greater resistance to excitotoxic cell death contributes to the high ammonia-tolerance of the goldfish. Whole-cell recordings measured in isolated brain slices of goldfish telencephalon during in vitro exposure to 5 mmol l(-1) or 10 mmol l(-1) T(Amm) reversibly potentiated NMDA receptor currents. This observation suggested that goldfish neurons may not be completely resistant to ammonia-induced excitotoxicity. Subsequent western blot and densitometric analyses revealed that NMDA receptor NR1 subunit abundance was 40-60% lower in goldfish exposed to 3-5 mmol l(-1) T(Amm) for 5 days, which was followed by a restoration of NR1 subunit abundance after 3 days recovery in ammonia-free water. We conclude that the goldfish brain may be protected from excitotoxicity by downregulating the abundance of functional NMDA receptors during periods when it experiences increased internal ammonia.
authors: Department of Biology, Wilfrid Laurier University, Waterloo, ON, Canada. mwilkie@wlu.ca
authors: Michael P
authors: MP
authors: Wilkie
authors: Matthew E
authors: ME
authors: Pamenter
authors: Stephanie
authors: S
authors: Duquette
authors: Hadi
authors: H
authors: Dhiyebi
authors: Navjeet
authors: N
authors: Sangha
authors: Geoffrey
authors: G
authors: Skelton
authors: Matthew D
authors: MD
authors: Smith
authors: Leslie T
authors: LT
authors: Buck
mesh_terms: D000641:Ammonia / Q000378:metabolism / Q000633:toxicity*; D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism; D016291:Dizocilpine Maleate / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005973:Glutamine / Q000378:metabolism; D006054:Goldfish / Q000378:metabolism*; D000860:Hypoxia / Q000378:metabolism*; D009584:Nitrogen / Q000097:blood / Q000378:metabolism; D017686:Oncorhynchus mykiss / Q000378:metabolism; D018408:Patch-Clamp Techniques; D016194:Receptors, N-Methyl-D-Aspartate / Q000037:antagonists & inhibitors / Q000378:metabolism*; D023382:Toxicity Tests, Acute; D014508:Urea / Q000378:metabolism; D014874:Water Pollutants, Chemical / Q000378:metabolism / Q000633:toxicity*
chemical_list: D018691:Excitatory Amino Acid Antagonists; D016194:Receptors, N-Methyl-D-Aspartate; D014874:Water Pollutants, Chemical; D005973:Glutamine; D016291:Dizocilpine Maleate; D000641:Ammonia; D014508:Urea; D009584:Nitrogen