Hypoxia transiently affects skeletal muscle hypertrophy in a functional overload model.

Hypoxia induces a loss of skeletal muscle mass, but the signaling pathways and molecular mechanisms involved remain poorly understood. We hypothesized that hypoxia could impair skeletal muscle hypertrophy induced by functional overload (Ov). To test this hypothesis, plantaris muscles were overloaded during 5, 12, and 56 days in female rats exposed to hypobaric hypoxia (5,500 m), and then, we examined the responses of specific signaling pathways involved in protein synthesis (Akt/mTOR) and breakdown (atrogenes). Hypoxia minimized the Ov-induced hypertrophy at days 5 and 12 but did not affect the hypertrophic response measured at day 56. Hypoxia early reduced the phosphorylation levels of mTOR and its downstream targets P70(S6K) and rpS6, but it did not affect the phosphorylation levels of Akt and 4E-BP1, in Ov muscles. The role played by specific inhibitors of mTOR, such as AMPK and hypoxia-induced factors (i.e., REDD1 and BNIP-3) was studied. REDD1 protein levels were reduced by overload and were not affected by hypoxia in Ov muscles, whereas AMPK was not activated by hypoxia. Although hypoxia significantly increased BNIP-3 mRNA levels at day 5, protein levels remained unaffected. The mRNA levels of the two atrogenes MURF1 and MAFbx were early increased by hypoxia in Ov muscles. In conclusion, hypoxia induced a transient alteration of muscle growth in this hypertrophic model, at least partly due to a specific impairment of the mTOR/P70(S6K) pathway, independently of Akt, by an undefined mechanism, and increased transcript levels for MURF1 and MAFbx that could contribute to stimulate the proteasomal proteolysis.

Control of gluconeogenic genes during intense/prolonged exercise: hormone-independent effect of muscle-derived IL-6 on hepatic tissue and PEPCK mRNA.

Prolonged intense exercise is challenging for the liver to maintain plasma glucose levels. Hormonal changes cannot fully account for exercise-induced hepatic glucose production (HGP). Contracting skeletal muscles release interleukin-6 (IL-6), a cytokine able to increase endogenous glucose production during exercise. However, whether this is attributable to a direct effect of IL-6 on liver remains unknown. Here, we studied hepatic glycogen, gluconeogenic genes, and IL-6 signaling in response to one bout of exhaustive running exercise in rats. To determine whether IL-6 can modulate gluconeogenic gene mRNA independently of exercise, we injected resting rats with recombinant IL-6. Exhaustive exercise resulted in a profound decrease in liver glycogen and an increase in gluconeogenic gene mRNA levels, phosphoenolpyruvate-carboxykinase (PEPCK), glucose-6-phosphatase (G6P), and peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha), suggesting a key role for gluconeogenesis in hepatic glucose production. This was associated to an active IL-6 signaling in liver tissue, as shown by signal transducer and activator of transcription and CAAT/enhancer binding protein-beta phosphorylation and IL-6-responsive gene mRNA levels at the end of exercise. Recombinant IL-6 injection resulted in an increase in IL-6-responsive gene mRNA levels in the liver. We found a dose-dependent increase in PEPCK gene mRNA strongly correlated with IL-6-induced gene mRNA levels. No changes in G6P and PGC-1alpha mRNA levels were found. Taken together, our results suggest that, during very demanding exercise, muscle-derived IL-6 could help increase HGP by directly upregulating PEPCK mRNA abundance.

Cyclosporin A inhibits hypoxia-induced pulmonary hypertension and right ventricle hypertrophy.

RATIONALE: Hypoxia-induced pulmonary hypertension involves hypoxia-inducible factor-1alpha (HIF-1alpha) activation as well as elevated resting calcium levels. Cyclosporin A (CsA) inhibits calcium-induced calcineurin activation and blocks the stabilization of HIF-1alpha in cultured cells. OBJECTIVES: We hypothesized that treatment of rats with CsA would prevent HIF-1-dependent gene transcription, lower specific responses to acute hypoxia, and prevent pulmonary hypertension and right ventricle hypertrophy resulting from prolonged exposure to hypoxia. METHODS: Acute and chronic responses to hypoxia were studied in rats treated or not treated with CsA (25 mg x kg(-1) x d(-1)). MEASUREMENTS: Transcript levels of genes encoding the serotonin transporter or four HIF-1 target genes, in rats exposed for 6 h to ambient hypoxia, treated or not by CsA, were measured. In vivo hemodynamics, hematocrit, and heart morphologic characteristics were assessed in rats subjected to hypoxia for 3 wk, treated or not treated with CsA. Changes in mRNA levels of the modulatory calcineurin-interacting protein-1 (MCIP-1) were used as a sensitive indicator of calcineurin activity in lung and heart. MAIN RESULTS: Acute exposure to hypoxia led to a marked increase in mRNA levels of serotonin transporter, modulatory calcineurin-interacting protein-1, and HIF-1 target genes, which was blunted by CsA treatment. Prolonged exposure to hypoxia raised right ventricle pressure, induced right ventricle hypertrophy, and activated cardiac calcineurin, effects that were fully prevented by CsA treatment. CONCLUSIONS: These results suggest that CsA prevents hypoxia-induced pulmonary hypertension and right ventricle hypertrophy, either by inhibiting HIF-1 transcriptional activity in lung, by decreasing calcineurin activity in lung and heart, by direct effects of CsA, or by a combination of these factors.

Leptin receptor-deficient obese Zucker rats reduce their food intake in response to hypobaric hypoxia.

Exposure to hypoxia induces anorexia in humans and rodents, but the role of leptin remains under discussion and that of orexigenic and anorexigenic hypothalamic neuropeptides remains unknown. The present study was designed to address this issue by using obese (Lepr(fa)/Lepr(fa)) Zucker rats, a rat model of genetic leptin receptor deficiency. Homozygous lean (Lepr(FA)/Lepr(FA)) and obese (Lepr(fa)/Lepr(fa)) rats were randomly assigned to two groups, either kept at ambient pressure or exposed to hypobaric hypoxia for 1, 2, or 4 days (barometric pressure, 505 hPa). Food intake and body weight were recorded throughout the experiment. The expression of leptin and vascular endothelial growth factor (VEGF) genes was studied in adipose tissue with real-time quantitative PCR and that of selected orexigenic and anorexigenic neuropeptides was measured in the hypothalamus. Lean and obese rats exhibited a similar hypophagia (38 and 67% of initial values at day 1, respectively, P < 0.01) and initial decrease in body weight during hypoxia exposure. Hypoxia led to increased plasma leptin levels only in obese rats. This resulted from increased leptin gene expression in adipose tissue in response to hypoxia, in association with enhanced VEGF gene expression. Increased hypothalamic neuropeptide Y levels in lean rats 2 days after hypoxia exposure contributed to accounting for the enhanced food consumption. No significant changes occurred in the expression of other hypothalamic neuropeptides involved in the control of food intake. This study demonstrates unequivocally that altitude-induced anorexia cannot be ascribed to anorectic signals triggered by enhanced leptin production or alterations of hypothalamic neuropeptides involved in anabolic or catabolic pathways.

Fibre-type specificity of interleukin-6 gene transcription during muscle contraction in rat: association with calcineurin activity.

In this study, we quantified the transcription of the interleukin-6 (IL-6) gene in individual fibres and the associated changes in calcineurin activity assessed at the cellular level during prolonged muscle contraction. Individual myofibres were isolated from plantaris and soleus muscles of rats at the end of an exhaustive running exercise test (n = 10), categorized according to their myosin heavy chain isoform content, and compared to those of resting rats (n = 10). Using real-time PCR analysis in individual fibres, a marked rise in IL-6 transcript levels occurred in type I and IIa fibres at the end of exercise (P < 0.05). Transcription of the gene encoding for the modulatory calcineurin-interacting protein-1 (MCIP-1), a sensitive indicator of calcineurin activity, also mainly increased in type I and IIa fibres (P < 0.05). Moreover, a slight increase in MCIP-1 mRNA levels was observed in type IIx (P < 0.05). Fibre types determined by immunohistochemistry were qualitatively examined for glycogen content using periodic acid-Shiff staining, and no direct relationship was found, at the cellular level, between glycogen content, fibre-type and IL-6 transcription. Our data clearly suggest that IL-6 gene transcription was mainly observed in early recruited myofibres and that contraction-induced IL-6 transcription could be associated with enhanced calcineurin activity.

Vascular endothelial growth factor expression in heart of rats exposed to hypobaric hypoxia: differential response between mRNA and protein.

This study investigated the role of vascular endothelial growth factor (VEGF) in the neoangiogenesis induced in heart in response to hypoxia. The time-course of adaptive changes in capillary supply, expression of VEGF mRNA and protein was studied in right (RV) and left ventricles (LV) of rats exposed to hypobaric hypoxia during 1-25 days (barometric pressure = 505 hPa). VEGF mRNA levels encoding for VEGF 188 and 164 isoforms were measured by reverse transcription-polymerase chain reaction (RT-PCR) and VEGF protein was determined by Western blotting. Relative RV weight (i.e., weight of RV related to body weight) increased with hypoxia and was 102% higher than in controls after 15 days of exposure (P < 0.01), while relative LV weight remained unaltered. A rapid and transient increase in VEGF 188 mRNA occurred after 1 day of hypoxia in LV (P < 0.05). Thereafter, a delayed increase in VEGF 188 mRNA expression occurred in RV (ANOVA, P < 0.001). By day 18, VEGF 188 mRNA level was higher in hypoxic than in control rats (P < 0.005) and then decreased to base line levels. Hypoxia did not affect the expression of VEGF 164 mRNA neither in LV nor in RV. One of the main results was that these hypoxia-induced alterations in VEGF transcripts were not followed by associated increase in VEGF protein. These results suggest that capillary growth observed in RV after prolonged exposure to ambient hypoxia likely results from other molecular mechanisms than VEGF.

MK801 impairs thermoregulation in the heat.

The effects of MK801 (dizocilpine), a glutamate NMDA receptor antagonist, on thermoregulation in the heat were studied in awake rats exposed to 40 degrees C ambient temperature until their body core temperature reached 43 degrees C. Under these conditions, MK801-treated rats exhibited enhanced locomotor activity and a steady rise in body core temperature, which reduced the heat exposure duration required to reach 43 degrees C. Since MK801-treated rats also showed increased striatal dopaminergic metabolism at thermoneutrality, the role of dopamine in the MK801-induced impairment of thermoregulation in the heat was determined using co-treatment with SCH23390, a dopamine D1 receptor antagonist. SCH23390 normalized the locomotor activity in the heat without any effect on the heat exposure duration. These results suggest that the MK801-induced impairment of thermoregulation in the heat is related to neither a dopamine metabolism alteration nor a locomotor activity enhancement.