Without oxygen, most vertebrates die within minutes as they cannot meet cellular energy demands with anaerobic metabolism. However, fish of the genus Carassius (crucian carp and goldfish) have evolved a specialized metabolic system that allows them to survive prolonged periods without oxygen by producing ethanol as their metabolic end-product. Here we show that this has been made possible by the evolution of a pyruvate decarboxylase, analogous to that in brewer’s yeast and the first described in vertebrates, in addition to a specialized alcohol dehydrogenase. Whole-genome duplication events have provided additional gene copies of the pyruvate dehydrogenase multienzyme complex that have evolved into a pyruvate decarboxylase, while other copies retained the essential function of the parent enzymes. We reveal the key molecular substitution in duplicated pyruvate dehydrogenase genes that underpins one of the most extreme hypoxic survival strategies among vertebrates and that is highly deleterious in humans.

Determination of muscle fiber composition in human skeletal muscle biopsies is often performed using immunohistochemistry, a method that tends to be both time consuming, technically challenging, and complicated by limited availability of tissue. Here, we introduce quantitative reverse transcriptase polymerase chain reaction (qRT-PCR)-based Gene-family profiling (GeneFam) of myosin heavy chain (MyHC) mRNA expression as a high-throughput, sensitive, and reliable alternative. We show that GeneFam and immunohistochemistry result in similar disclosures of alterations in muscle fiber composition in biopsies from musculus vastus lateralis and musculus biceps brachii of previously untrained young women after 12 weeks of progressive strength training. The adaptations were evident as (a) consistent increases in MyHC2A abundance; (b) consistent decreases in MyHC2X abundance; and (c) consistently stable MyHC1 abundance, and were not found using traditional reference gene-based qRT-PCR analyses. Furthermore, muscle fiber composition found using each of the two approaches was correlated with each other (r = 0.50, 0.74, and 0.78 for MyHC1, A, and X, respectively), suggesting that GeneFam may be suitable for ranking of individual muscle phenotype, particularly for MyHC2 fibers. In summary, GeneFam of MyHC mRNA resulted in reliable assessment of alterations in muscle fiber composition in skeletal muscle of previously untrained women after 12 weeks of strength training.

Ellefsen S, Stensløkken K, Fagernes CE, Kristensen TA, Nilsson GE. Expression of genes involved in GABAergic neurotransmission in anoxic crucian carp brain (Carassius carassius). Physiol Genomics 36: 61-68, 2009. First published October 28, 2008 doi:10.1152/physiolgenomics.90301.2008.-The crucian carp, Carassius carassius, survives days to months without oxygen, depending on temperature. In the anoxic crucian carp brain, increased GABAergic inhibition, mediated by increased extracellular levels of GABA, has been shown to suppress electric activity and ATP consumption. To investigate an involvement of gene expression in this response, we utilized real-time RT-PCR to test the effect of 1 and 7 days anoxia (8°C) on the expression of 22 genes, including nine GABAA receptor subunits (␣1-6, ␤2, ␦, and ␥2), three GABAB receptor subunits (GB1a-1b and GB2), three enzymes involved in GABA metabolism (GAD65 and GAD67, GABAT), four GABA transporters (GAT1, 2a-b and 3), two GABAA receptor-associated proteins (GABARAP 1 and 2), and the K ϩ /Cl Ϫ cotransporter KCC2. While the expression of GABAA receptor subunits was dominated by ␣4-, ␣6-, and ␦-subunits, all of which are located to extrasynaptic sites in mammalian brains and respond to elevations in extracellular levels of GABA by showing tonic activity patterns, the expression of GABA transporters was dominated by GAT2 (a and b) and GAT3, which also show extrasynaptic location in mammals. These expression patterns differ from those observed in mammals and may be a prerequisite for GABAergic inhibition of anoxic metabolic rate in crucian carp. Furthermore, while the expression of the majority of the genes was largely unaltered by anoxia, the expression of GAT2 and GAT3 decreased to 20%. This suggests impairment of GABA transport, which could be a mechanism behind the accumulation of extracellular GABA and the increased GABAergic inhibition.GABA; GABA receptor; GABA transporter; external RNA control SINCE GABA IS THE DOMINANT inhibitory neurotransmitter in vertebrate brains, GABAergic neurotransmission has been suggested to function as an endogenous defense against excitotoxic events, such as the excessive release of glutamate during periods of ischemia/anoxia (26). However, GABAergic activity seems to be lowered rather than enhanced in ischemic mammalian brains, resulting in neuronal hyperexcitability and cell death (41). Thus, efforts have been made to reverse this process, and several studies have found that artificially increased GABAergic activity results in increased neuroprotection (41).Interestingly, anoxia induces increased GABA levels and GABAergic activity in brains of vertebrate species that tolerate prolonged periods of oxygen depletion, such as fish of the genus Carassius and freshwater turtles of genera Chrysemys and Trachemys. For example, while crucian carp (Carassius carassius) brains show a doubling of extracellular GABA levels after 6 h of anoxia and a fivefold increase in tissue GABA levels after prolonged anoxia (32, 19), freshwater turtle brains show a ...

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We report here the first comprehensive gene expression analysis of GABAA receptor subunits and of genes involved in brain GABAergic transmission in fish exposed to near-future CO2 conditions. Altogether, 56 mRNA transcripts were quantified in brains of three-spined stickleback (Gasterosteus aculeatus) kept in control or elevated pCO2 for 43 days.

The continuous increase of anthropogenic CO in the atmosphere resulting in ocean acidification has been reported to affect brain function in some fishes. During adulthood, cell proliferation is fundamental for fish brain growth and for it to adapt in response to external stimuli, such as environmental changes. Here we report the first expression study of genes regulating neurogenesis and neuroplasticity in brains of three-spined stickleback (), cinnamon anemonefish () and spiny damselfish () exposed to elevated CO The mRNA expression levels of the neurogenic differentiation factor (NeuroD) and doublecortin (DCX) were upregulated in three-spined stickleback exposed to high-CO compared with controls, while no changes were detected in the other species. The mRNA expression levels of the proliferating cell nuclear antigen (PCNA) and the brain-derived neurotrophic factor (BDNF) remained unaffected in the high-CO exposed groups compared to the control in all three species. These results indicate a species-specific regulation of genes involved in neurogenesis in response to elevated ambient CO levels. The higher expression of NeuroD and DCX mRNA transcripts in the brain of high-CO-exposed three-spined stickleback, together with the lack of effects on mRNA levels in cinnamon anemonefish and spiny damselfish, indicate differences in coping mechanisms among fish in response to the predicted-future CO level.

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