Stress and interpopulation variation in glycolytic enzyme activity and expression in a teleost fish Fundulus heteroclitus

Northern populations of Fundulus heteroclitus have twofold greater activity of lactate dehydrogenase-B (LDH-B) than southern populations, but exposure to stress increases LDH-B in southern populations, abolishing this difference. To test whether differences in the activity of other hepatic glycolytic enzymes between populations are sensitive to stress, we injected fish with a pharmacological dose of cortisol in coconut oil (400 microg g(-1)) or exposed them to handling stress and measured the activities of all the glycolytic enzymes. At rest, liver phosphofructokinase (PFK) and aldolase (ALD) activities were greater in southern fish, whereas LDH-B activity was greater in northern fish. No other glycolytic enzymes differed in activity between populations in control fish. Cortisol injection and handling stress decreased PFK and ALD and increased LDH activities in the southern but not the northern population, such that the populations no longer differed in the activity of any enzyme following treatment. Unlike Ldh-B mRNA, Pfk and Ald mRNA levels did not parallel enzyme activity, suggesting complex kinetics or regulation at multiple levels. Plasma cortisol did not differ between populations at rest but was significantly different between populations in treated fish. These data suggest that differences in liver enzyme activity may be related to differences in stress hormone physiology between populations.     

Ontogenetic changes and developmental adjustments in lactate dehydrogenase isozymes of an obligate air-breathing fish Channa punctatus during deprivation of air access

In air-breathing snakehead Channa punctatus, Ldh-B is expressed at all ontogenetic and developmental stages, while Ldh-A is expressed temporally in pre-hatchlings 12-13 days ahead of bimodal respiration marked by air-breathing. Remarkable differences are observed in the LDH isozyme expression among various ontogenetic and developmental stages upon denying air access. When denied air access, water-breathing larvae show two distinct characteristics: (i) they survive longer than transitory air-breathers due to independence from air-breathing and (ii) there is more transient induction of Ldh-B than Ldh-A. Transition to bimodal breathing, which occurred post-hatching in 15-day old larvae, is coincidental with inducibility of Ldh-A and concomitant down-regulation of Ldh-B. Heart tissue from air-breathing adults denied air access shows a preferential expression of LDH-A subunit and slight down-regulation of LDH-B. Heterotetramers of A and B subunits participate in adjusting LDH levels among those stages which either precede air-breathing switchover, or are subsequent to this transition. The contribution of heterotetramers depends on the stage-specific levels of LDH homotetramers A(4) or B(4). Scaling of muscle mass during growth, tolerance to extended deprivation of air access and induction of Ldh-A are correlated. Response to restoring air contact indicated that advanced air-breathing stages of C. punctatus possess an inherent capacity to sense surface air. In kinetic properties, LDH isozymes of C. punctatus are teleost-like but species specificity is displayed in oxidative potential by cardiac muscle and in L-lactate reduction by skeletal muscle.     

Lactate dehydrogenase expression at the onset of altered loading in rat soleus muscle.

Both functional overload and hindlimb disuse induce significant energy-dependent remodeling of skeletal muscle. Lactate dehydrogenase (LDH), an important enzyme involved in anaerobic glycolysis, catalyzes the interconversion of lactate and pyruvate critical for meeting rapid high-energy demands. The purpose of this study was to determine rat soleus LDH-A and -B isoform expression, mRNA abundance, and enzymatic activity at the onset of increased or decreased loading in the rat soleus muscle. The soleus muscles from male Sprague-Dawley rats were functionally overloaded for up to 3 days by a modified synergist ablation or subjected to disuse by hindlimb suspension for 3 days. LDH mRNA concentration was determined by Northern blotting, LDH protein isoenzyme composition was determined by zymogram analysis, and LDH enzymatic activity was determined spectrophotometrically. LDH-A mRNA abundance increased by 372%, and LDH-B mRNA abundance decreased by 43 and 31% after 24 h and 3 days of functional overload, respectively, compared with that in control rats. LDH protein expression demonstrated a shift by decreasing LDH-B isoforms and increasing LDH-A isoforms. LDH-B activity decreased 80% after 3 days of functional overload. Additionally, LDH-A activity increased by 234% following 3 days of hindlimb suspension. However, neither LDH-A or LDH-B mRNA abundance was affected following 3 days of hindlimb suspension. In summary, the onset of altered loading induced a differential expression of LDH-A and -B in the rat soleus muscle, favoring rapid energy production. Long-term altered loading is associated with myofiber conversion; however, the rapid changes in LDH at the onset of altered loading may be involved in other physiological processes.     

Differential performance among LDH-B genotypes in Rana lessonae tadpoles

The European pool frog, Rana lessonae, is widely polymorphic for two common alleles (b,e) at the lactate dehydrogenase-B (LDH-B) locus. We compared fitness-related larval life-history traits among LDH-B genotypes, which originated from segregation in heterozygous parents, in an artificial pond experiment where tadpoles of R. lessonae from a Swiss population were raised together with tadpoles of the hemiclonal hybrid R. esculenta at two densities. In R. lessonae, LDH-B e/e homozygotes at each density had a higher proportion of metamorphs among survivors, reached metamorphosis earlier, and were heavier at metamorphosis than b/b homozygotes; b/e heterozygotes had intermediate values. That e/e individuals were superior to b/b in both time to and mass at metamorphosis is surprising because these two life-history traits are thought to reflect a performance trade-off; e/e genotypes apparently compensated for shorter time to metamorphosis by a higher growth rate. The two alleles showed the same performance ranking when combined in hybrids with a R. ridibunda allele: When R. esculenta from Swiss populations reared in the same ponds had received the e allele rather than the b allele from their R. lessonae parent, they reached metamorphosis earlier, but did not differ in mass at metamorphosis. The degree of linkage disequilibrium in the source population of the eight R. lessonae used as parents of the R. lessonae tadpoles is unknown, so we cannot exclude the possibility that the performance differences are caused by some anonymous tightly linked gene, rather than the LDH-B locus, that constitutes the genomically localized target of natural selection. A causal involvement of LDH-B is plausible, nevertheless, because this enzyme takes part in the central energy-metabolizing processes and has been reported to underlie fitness differences in other animals; also, differential performance of LDH-B genotypes has been observed in R. lessonae larvae from another population. The present results suggest strong directional selection for allele e; the sum of available data, including an independent laboratory experiment, suggests that partial environment-dependent overdominance combined with balancing selection favoring e/e homozygotes under some and b/b homozygotes under other conditions may be partially responsible for the broad maintenance of the LDH-B polymorphism in R. lessonae.     

Gene expression during mammalian oogenesis and early embryogenesis: quantification of three messenger RNAs abundant in fully grown mouse oocytes

Ribonuclease protection assays have been used to quantitatively assess changes in steady-state levels of specific mRNAs during oogenesis and early embryogenesis in mice. The mRNAs encode ZP3 (a glycoprotein that serves as a sperm receptor), LDH-B (heart-type lactate dehydrogenase), and MOM-1 (a protein of unknown function). MOM-1 and LDH-B are expressed in a variety of adult mouse tissues and midgestation embryos, whereas ZP3 expression is restricted completely to oocytes. All three mRNAs are expressed by growing mouse oocytes and accumulate to unusually high levels in fully grown oocytes as compared to somatic cells; 240,000, 200,000 and 74,000 copies mRNA per fully grown oocyte for ZP3, LDH-B and MOM-1, respectively. Steady-state levels of LDH-B and MOM-1 mRNA undergo a modest decline (approximately 20-40%) during ovulation when fully grown oocytes become unfertilized eggs and, in general, mirror the reported change in poly(A)+RNA levels during this period of development. On the other hand, the level of ZP3 mRNA declines dramatically (approximately 98%) during ovulation, from approximately 240,000 copies per oocyte to approximately 5000 copies per unfertilized egg, and ZP3 mRNA is undetectable in fertilized eggs (less than 1000 copies per fertilized egg). MOM-1 mRNA is expressed at relatively low levels in morulae (approximately 2000 copies per embryo) and blastocysts (approximately 5000 copies per embryo), whereas ZP3 mRNA remains undetectable (less than 1000 copies per embryo) at these stages of preimplantation development. These findings are discussed in the context of overall gene expression during oocyte growth, meiotic maturation and early embryogenesis in mice.     

A Mutation Affecting the Lactate Dehydrogenase Locus Ldh-1 in the Mouse. II. Mechanism of the Ldh-a Deficiency Associated with Hemolytic Anemia

A procarbazine hydrochloride-induced mutation at the Ldh-1 structural locus encoding the A subunit of lactate dehydrogenase (LDH) was used to study the molecular and metabolic basis of severe hemolytic anemia due to LDH-A deficiency in the mouse. The mutant allele designated Ldh-1(a-m1Neu) codes for an enzyme that as homotetramer differs from the wild-type enzyme by a marked instability, acidic shift of the pH profile, increased K(m) for pyruvate and altered inhibition by high concentrations of this substrate. Except for the latter, all these altered properties of the mutant protein contribute to the diminished LDH activity in heterozygous and homozygous mutant individuals. Impaired energy metabolism of erythrocytes indicated by a relatively low ATP concentration is suggested to result in cell death at the end of the reticulocyte stage leading to the expression of hemolytic anemia with extreme reticulocytosis and hyperbilirubinemia. Despite the severe anemia, affected homozygous mutants exhibit approximately normal body weight and do not show noticeable impairment of viability or fertility. To date no such condition is observed in man. This discrepancy is likely due to the fact that in human erythrocytes both LDH-A and LDH-B subunits are expressed such that homozygotes for a LDH-A or LDH-B deficiency would not result in a comparably extreme LDH activity deficiency.