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A key determinant of geriatric frailty is sarcopenia, the age-associated loss of skeletal muscle mass and strength (1,2). Although the etiology of sarcopenia is unknown, the correlation during aging between the loss of activity of satellite cells, which are endogenous muscle stem cells, and impaired muscle regenerative capacity has led to the hypothesis that the loss of satellite cell activity is also a cause of sarcopenia (3,4). We tested this hypothesis in male sedentary mice by experimentally depleting satellite cells in young adult animals to a degree sufficient to impair regeneration throughout the rest of their lives. A detailed analysis of multiple muscles harvested at various time points during aging in different cohorts of these mice showed that the muscles were of normal size, despite low regenerative capacity, but did have increased fibrosis. These results suggest that lifelong reduction of satellite cells neither accelerated nor exacerbated sarcopenia and that satellite cells did not contribute to the maintenance of muscle size or fiber type composition during aging, but that their loss may contribute to age-related muscle fibrosis., Recent estimates indicate that up to one-third of elderly people suffer from frailty, characterized by a common set of symptoms including loss of muscle strength, increased fatigability, modest levels of [...]

MyoD, a master regulator of myogenesis, exhibits a circadian rhythm in its mRNA and protein levels, suggesting a possible role in the daily maintenance of muscle phenotype and function. We report that MyoD is a direct target of the circadian transcriptional activators CLOCK and BMAL1, which bind in a rhythmic manner to the core enhancer of the MyoD promoter. Skeletal muscle of [Clock.sup.[DELTA]19] and [Bmal1.sup.-/-] mutant mice exhibited ~30% reductions in normalized maximal force. A similar reduction in force was observed at the single-fiber level. Electron microscopy (EM) showed that the myofilament architecture was disrupted in skeletal muscle of [Clock.sup.[DELTA]19], [Bmal.sup.1-/-], and [MyoD.sup.-/-] mice. The alteration in myofilament organization was associated with decreased expression of actin, myosins, titin, and several MyoD target genes. EM analysis also demonstrated that muscle from both [Clock.sup.[DELTA]19] and [Bmal1.sup.-/-] mice had a 40% reduction in mitochondrial volume. The remaining mitochondria in these mutant mice displayed aberrant morphology and increased uncoupling of respiration. This mitochondrial pathology was not seen in muscle of [MyoD.sup.-/-] mice. We suggest that altered expression of both Pgc-1[alpha] and Pgc-1[beta] in [Clock.sup.[DELTA]19] and [Bmal1.sup.-/-] mice may underlie this pathology. Taken together, our results demonstrate that disruption of CLOCK or BMAL1 leads to structural and functional alterations at the cellular level in skeletal muscle. The identification of MyoD as a clock-controlled gene provides a mechanism by which the circadian clock may generate a muscle-specific circadian transcriptome in an adaptive role for the daily maintenance of adult skeletal muscle. circadian clock | myofilaments | mitochondria doi/ 10.1073/pnas.1014523107

Sarcopenia, skeletal muscle loss during aging, is associated with increased falls, fractures, morbidity, and loss of independence. MicroRNAs (miRNAs) are novel posttranscriptional regulators. The role of miRNAs in cell size regulation after an anabolic stimulus in human skeletal muscle is unknown. We hypothesized that aging would be associated with a differential expression of skeletal muscle primary miRNA (pri-miRNA) and mature miRNA (miR). To test this hypothesis, we used real-time PCR and immunoblotting before and after an anabolic stimulus (resistance exercise + ingestion of a 20-g leucine-enriched essential amino acid solution) to measure the expression of muscle-specific miRNAs (miR-1, miR-133a, and miR-206), upstream regulators (MyoD and myogenin), and downstream targets [insulin-like growth factor-I, histone deacetylase-4, myocyte enhancing factor-2, and Ras homolog enriched in brain (Rheb)] in skeletal muscle of young and older men. Muscle biopsies were obtained at baseline and 3 and 6 h after exercise. At baseline, we found pri-miRNA-1-1, -1-2, -133a-1, and -133a-2 expression elevated in older compared with young men (P < 0.05). Pri-miRNA-1-2, -133a-1, and -133a-2 were reduced at 6 h after exercise only in the young men compared with baseline, whereas pri-miRNA-206 was elevated at different postexercise time points in older and young men (P < 0.05). Compared with baseline, miR-1 was reduced only in the young men, whereas Rheb protein was increased in both age groups after the anabolic stimulus (P < 0.05). We conclude that skeletal muscle primary and mature miRNA expression in young men is readily altered by an anabolic stimulus of resistance exercise + essential amino acid ingestion. However, aging is associated with higher basal skeletal muscle primary miRNA expression and a dysregulated miRNA response after the anabolic stimulus. muscle growth; sarcopenia; posttranscriptional regulators

To assess potential risks related to the duration or total amount of fetal methadone exposure during gestation, we compared babies of women who conceived and maintained on methadone throughout pregnancy with babies of women who began methadone treatment during the second or third trimester. Babies conceived on methadone were exposed to the medication for a mean of 37.4 weeks at a mean dose of 110 mg/day, whereas comparison babies were exposed for a mean of 13.1 weeks at a mean dose of 93 rag/day. There were no significant between-group differences in the frequency of treatment of neonatal abstinence, days hospitalized, birth weight, or gestational age. Babies conceived on methadone were significantly less likely to test positive for illicit drugs at delivery as compared with babies conceived off methadone (positive toxicology, 9.1% vs. 34.3%, respectively). Methadone exposure during the entire gestational period was associated with better drug-treatment outcomes but was not associated with more severe neonatal abstinence. Keywords: Methadone; Pregnancy; Conception; Neonatal abstinence; Drug treatment

Neurons of the organum vasculosum of the lamina terminalis (OVLT) are necessary for thirst and vasopressin secretion during hypersmolality in rodents. Recent evidence suggests the osmosensitivity of these neurons is mediated by a gene product encoding the transient receptor potential vanilloid- 1 (TRPV1) channel. The purpose of the present study was to determine whether mice lacking the TRPV1 channel had blunted thirst responses and central Fos activation to acute and chronic hyperosmotic stimuli. Surprisingly, TRPV1-/- vs. wild-type mice ingested similar amounts of water after injection (0.5 ml sc) of 0.5 M NaCl and 1.0 M NaCl. Chronic increases in plasma osmolality produced by overnight water deprivation or sole access to a 2% NaCl solution for 48 h produced similar increases in water intake between wild-type and TRPVI-/- mice. There were no differences in cumulative water intakes in response to hypovolemia or isoproterenol. In addition, the number of Fos-positive cells along the lamina terminalis, including the OVLT, as well as the supraoptic nucleus and hypothalamic paraventricular nucleus, was similar between wild-type and TRPV1-/- mice after both acute and chronic osmotic stimulation. These findings indicate that TRPV1 channels are not necessary for osmotically driven thirst or central Fos activation, and thereby suggest that TRPV1 channels are not the primary ion channels that permit the brain to detect changes in plasma sodium concentration or osmolality. water intake; vasopressin; oxytocin; organum vasculosum of the lamina terminalis; osmolality

MicroRNAs are highly conserved, noncoding RNAs involved in posttranscriptional gene silencing. MicroRNAs have been shown to be involved in a range of biological processes, including myogenesis and muscle regeneration. The objective of this study was to test the hypothesis that microRNA expression is altered in dystrophic muscle, with the greatest change occurring, of the muscles examined, in the diaphragm. The expression of the muscle-enriched microRNAs was determined in the soleus, plantaris, and diaphragm muscles of control and dystrophin-deficient (mdx) mice by semiquantitative PCR. In the soleus and plantaris, expression of the mature microRNA 133a (miR-133a) and miR-206, respectively, was decreased by ~25%, whereas in the diaphragm, miR-206 expression increased by 4.5-fold relative to control. The increased expression of miR-206 in the mdx diaphragm was paralleled by a 4.4-fold increase in primary miRNA-206 (primiRNA-206) transcript level. Expression of Myodl was elevated 2.7-fold only in the mdx diaphragm, consistent with an earlier finding demonstrating Myodl can activate pri-miRNA-206 transcription. Transcript levels of Drosha and Dicer, major components of microRNA biogenesis pathway, were unchanged in mdx muscle, suggesting the pathway is not altered under dystrophic conditions. Previous in vitro analysis found miR-206 was capable of repressing utrophin expression; however, under dystrophic conditions, both utrophin transcript and protein levels were significantly increased by 69% and 3.9-fold, respectively, a finding inconsistent with microRNA regulation. These results are the first to report alterations in expression of muscle-enriched microRNAs in skeletal muscle of the mdx mouse, suggesting microRNAs may have a role in the pathophysiology of muscular dystrophy. muscular dystrophy; posttranscriptional; gene regulation doi:10.1152.ajpcell.00077.2007

The initial asymmetry that specifies the oral-aboral (OA) axis of the sea urchin embryo has long been a mystery. It was shown previously that OA polarity can be entrained in embryos by imposing a respiratory asymmetry, with the most oxidizing side of the embryo tending to develop as the oral pole. This suggests that one of the earliest observable asymmetries along the incipient OA axis, a redox gradient established by a higher density and/or activity of mitochondria on the prospective oral side of the embryo, might play a causal role in establishing the axis. Here, we examine the origin and functional significance of this early redox gradient. Using MitoTracker Green, we show that mitochondria are asymmetrically distributed in the unfertilized egg of Strongylocentrotus purpuratus, and that the polarity of the maternal asymmetry is maintained in the zygote. Vital staining indicates that the side of the embryo that inherits the highest density of mitochondria tends to develop into the oral pole. This correlation holds when mitochondria are redistributed by centrifugation of eggs or by transfer of purified mitochondria into zygotes, indicating that an asymmetric mitochondrial distribution can entrain OA polarity, possibly through effects on intracellular redox state. In support of this possibility, we find that specification of oral ectoderm is suppressed when embryos are cultured under hypoxic conditions that enforce a relatively reducing redox state. This effect is reversed by overexpression of nodal, an early zygotic marker of oral specification whose localized expression suffices to organize the entire OA axis, indicating that redox state is upstream of nodal expression. We therefore propose that a threshold level of intracellular oxidation is required to effectively activate nodal, and that precocious attainment of this threshold within the blastomeres containing the highest density of mitochondria results in asymmetric nodal activity and consequent specification of the OA axis. Keywords: Sea urchin; Development; Mitochondria; Redox state; Transcription; Metabolic gradient

We revised distribution maps of potential presettlement habitat and current populations for Greater Sage-Grouse (Centrocercus urophasianus) and Gunnison Sage-Grouse (C. minimus) in North America. The revised map of potential presettlement habitat included some areas omitted from previously published maps such as the San Luis Valley of Colorado and Jackson area of Wyoming. Area excluded from the revised maps were those dominated by barren, alpine, and forest habitats. The resulting presettlement distribution of potential habitat for Greater Sage-Grouse encompassed 1 200 483 [km.sup.2], with the species' current range 668 412 [km.sup.2]. The distribution of potential Gunnison Sage-Grouse habitat encompassed 46 521 [km.sup.2], with the current range 4787 [km.sup.2]. The dramatic differences between the potential presettlement and current distributions appear related to habitat alteration and degradation, including the adverse effects of cultivation, fragmentation, reduction of sagebrush and native herbaceous cover, development, introduction and expansion of invasive plant species, encroachment by trees, and issues related to livestock grazing. Key words: Centrocercus minimus, Centrocercus urophasianus, distribution, Greater Sage-Grouse, Gunnison Sage-Grouse, habitat change.

The mechanisms that induce the reduction of beta-myosin heavy chain (MHC) expression in response to non-weight bearing activity were analyzed in soleus muscles isolated from mice subjected to hindlimb suspension (HS). Northern blot analysis of HS mouse soleus indicated the presence of reduced levels of messenger RNA (mRNA) expression for beta-MHC, cardiac/slow troponin C and slow myosin light chain one and two. HS also increased the levels of muscle creatine kinase and skeletal troponin C mRNA expression.

Thee induction of beta-myosin heavy chain (beta-MHC) gene expression was investigated in transgenic mouse lines containing serial deletions of the human beta-MHC promoter to nucleotides -293, -201, and -141. Expression assays identified only the 293-base pair beta-MHC in both adult and fetal mice muscle. Expression of the 293 gene was site specific at the plantaris and soleus muscles.

Mutations of the DNA regulatory sub-regions, muscle chloramphenicol acetyltransferase (MCAT), C-rich and beta-e3, fail to eliminate the induction of beta-myosin heavy chain (MHC) transgene in mechanically overloaded plantaris muscle of transgenic mice. Protein extracts from these wild-type muscles and those containing a mutant transgene show a marked increase in CAT specific activity. In transgenic mice with mutations in a 5,600-base pair of the transgene, the increased CAT activity is restricted to type I MHC fibres.