In the rat diaphragm muscle, the histochemical classification of type I, IIa, IIb, or IIx fibers was correlated with myosin heavy chain (MHC) immunoreactivity. Expression of MHC isoforms in single dissected fibers was also assessed electrophoretically. Most fibers (approximately 86%) expressed a single MHC isoform, and when present, coexpression of MHC-2X and MHC-2B isoforms was most prevalent. Type I and IIa fibers were the smallest, type IIb fibers were the largest, and type IIx fibers were intermediate. Succinate dehydrogenase (SDH) and calcium-activated myosin adenosinetriphosphatase (actomyosin ATPase) activities were measured with quantitative histochemical procedures. Type I and IIa fibers had the highest SDH activities, followed in rank order by type IIx and IIb fibers. Type I fibers had the lowest actomyosin ATPase activity, followed in rank order by type IIa, IIx, and IIb fibers. Across all fibers, there was an inverse relationship between fiber SDH activity and cross-sectional area and a positive correlation between fiber actomyosin ATPase activity and cross-sectional area. The SDH and actomyosin ATPase activities of muscle fibers were also inversely correlated. These phenotypic differences in SDH and ATPase activities may be important in determining the contractile and fatigue properties of different fiber types in the rat diaphragm muscle.

P-glycoprotein (P-gp/ABCB1), multidrug resistance protein 1 (MRP1/ABCC1), and breast cancer resistance protein (BCRP/ABCG2) are plasma membrane efflux pumps that limit the intracellular uptake and retention of numerous lipophilic, amphipathic xeno-and endobiotics. Little is known about the neonatal and developmental expression of P-gp/ABCB1, MRP1/ABCC1, and BCRP/ ABCG2 in the human central nervous system (CNS), therefore postmortem CNS tissue from infants born 22 0/7 -42 0/7 week gestation and adults was immunostained to determine their ontogeny and cellular localization. P-gp/ABCB1 imunostaining was observed in microvessel endothelial cells as early as 22 0/7 weeks, increasing in prevalence and intensity with maturation, and later in gestation in large pyramidal neurons. MRP1/ABCC1 immunostaining was prominent early in the choroid plexus and ventricular ependyma, and noted later in large pyramidal neurons. BCRP/ABCG2 expression was limited to microvessel endothelial cells. P-gp/ABCB1, MRP1/ABCC1 and BCRP/ ABCG2 in adult brain matched term newborn CNS but with more intense immunostaining. We conclude that P-gp/ABCB1, MRP1/ABCC1, and BCRP/ABCG2 are expressed in a developmental, cell specific, fashion in the human CNS. The complementary pattern of P-gp/ABCB1 and BCRP/ ABCG2 at the blood-brain with MRP1/ABCC1 at the blood-CSF barriers may limit CNS uptake and retention of drugs and toxins in neonates.

The objective of this study was to determine the relationship between developmental transitions in myosin heavy chain (MHC) composition and changes in maximum unloaded shortening velocity (Vo) and maximum specific force (Po) of the rat diaphragm muscle. The diaphragm was excised at postnatal days 0, 3, 7, 14, 21, and 28 and in adults. MHC isoform expression was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and laser densitometry. In muscle fiber bundles, Vo was determined at 15 degrees C by use of the "slack" test. Isometric Po was determined at 15 and 26 degrees C. Simple and stepwise regressions were used to evaluate the correlations between Vo, Po, and MHC phenotype transitions and the various developmental ages. The progressive increases in Vo and Po with age were found to be inversely correlated to MHC-neonatal isoform expression (r2 = -0.84 and -0.63, respectively) and positively correlated to MHC-2X (r2 = 0.78 and 0.57) and MHC-2B (r2 = 0.51 and 0.40) isoform expression (P < 0.001). Changes in MHC-neonatal isoform expression contributed to most of the developmental variance in Vo and Po, with changes in MHC-2X and MHC-2B expression also contributing significant increments to total variance. The postnatal increase in Vo most likely relates to differences in the actomyosin adenosinetriphosphatase activity between neonatal and adult fast MHC phenotypes. The increase in Po may reflect inherent differences in myofibrillar density, cross-bridge cycling kinetics, and/or the force produced per cross bridge among fibers composed of the different MHC isoforms.

A profile of respiratory complications has been associated with the onset and development of obesity in humans. Similar phenotypes have been routinely demonstrated in genetic animal models of obesity such as the ob mouse (C57BL/6J-Lepob). The objective of the present study was to test the hypothesis that a constellation of respiratory complications are attenuated with leptin (i.e., protein product of the ob gene) replacement. Daily leptin administration during a 6-wk period was conducted to control body weight of mutant ob mice similar to genotypic control groups. During the treatment period, repeated baseline ventilatory measurements were assessed by using whole body plethysmography while quasistatic pressure-volume curves were performed to further explore the role of leptin in improving lung mechanics. Diaphragmatic myosin heavy chain (MHC) isoform phenotype was examined to determine proportional changes in MHC composition. In room air, breathing frequency and minute ventilation were significantly (P < 0.01) different among ob treatment groups, suggesting that leptin opposed the development of a rapid breathing pattern observed in vehicle-treated ob mice. Quasistatic deflation curves indicated that the lung volume of leptin-treated ob mice was significantly (P < 0.05) greater relative to vehicle-treated ob mice at airway pressures between 0 and 30 cmH2O. Diaphragm MHC composition of leptin-treated ob mice was restored significantly (P < 0.05) to resemble the control phenotype. In this genetic mouse model of obesity, the results suggested that respiratory complications associated with the obese phenotype, including rapid breathing pattern at baseline, diminished lung compliance, and abnormal respiratory muscle adaptations, are attenuated with prolonged leptin treatment.

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Uteroplacental insufficiency and subsequent intrauterine growth retardation (IUGR) increase the risk of insulin resistance in humans and rats. Aberrant skeletal muscle lipid metabolism contributes to the pathogenesis of insulin resistance. Peroxisome proliferator-activated receptor-␥ co-activator-1 (PGC-1) is a transcriptional co-activator that affects gene expression of key lipid metabolizing enzymes such as carnitine palmitoyltransferase I (mCPTI). Because gene expression of lipid metabolizing enzymes is altered in IUGR postnatal skeletal muscle, and we hypothesized that PGC-1 expression would be similarly affected. To prove this hypothesis, bilateral uterine artery ligation and sham surgery were used to produce IUGR and control rats respectively. Western Blotting demonstrated that PGC-1 hind limb skeletal muscle protein levels were increased in perinatal and postnatal IUGR rats. Conventional RT-PCR demonstrated that PGC-1 mRNA levels were similarly increased in perinatal hind limb skeletal muscle and juvenile extensor digitorum longus (EDL), but were decreased in juvenile soleus. Because a gender specific trend was noted in PGC-1 mRNA levels, real time RT-PCR was used for further differentiation. Real time RT-PCR revealed that changes in postnatal skeletal muscle PGC-1 expression were more marked in male IUGR rats versus female IUGR rats. Down stream targets of PGC-1 followed a similar pattern of expression. We conclude that PGC-1 expression is altered in rat IUGR skeletal muscle and speculate that it contributes to the pathogenesis of insulin resistance in the IUGR rat. Barker's Fetal Origins of Adult Disease Hypothesis proposes that fetal adaptation to a deprived intrauterine milieu leads to permanent changes in cellular biology and systemic physiology (1). Intrauterine growth retardation (IUGR) predisposes affected newborns toward the development of insulin resistance and dyslipidemia (2). Although both insulin deficiency and resistance contribute to the IUGR diabetic phenotype, asymmetrical IUGR individuals are often characterized by insulin resistance. Uteroplacental insufficiency, a morbidity associated with many common complications of pregnancy induces asymmetrical IUGR (3, 4). In the rat, uteroplacental insufficiency results in juvenile IUGR animals whose glucose homeostasis is abnormal only when physiologically challenged on a pharmacological level (5). By adulthood, these IUGR rats develop overt diabetes that is characterized by insulin resistance and hypertriglyceridemia (5,19,20).An important component contributing to the pathogenesis of skeletal muscle insulin resistance is altered fatty acid homeostasis (6, 7

The mdr1a isoform of P-glycoprotein (Pgp) is an integral plasma membrane efflux pump expressed in adult brain capillary endothelial cells and astrocytes of the blood-brain barrier. We determined the developmental pattern of Pgp expression in brain tissue at embryonic day 16 (E16), day of life 0 (D0), day of life 7 (D7), day of life 21 (D21), and adults (Ad). The relative expression of Pgp mRNA and protein was indexed as a percent (mean ± SEM) of D0 levels. Pgp mRNA levels increased significantly (p < 0.01) with maturation (E16: 75 ± 8%; D21: 303 ± 37%, and Ad: 1,160 ± 120%). Similarly, Pgp protein expression was observed at E16 and increased significantly (p < 0.01) during development (E16: 52 ± 8%; D7: 187 ± 23%; D21: 440 ± 48%, and Ad: 441 ± 56%). This developmental pattern of enhanced blood-brain barrier Pgp expression with maturation was confirmed by immunohistochemistry. We conclude that (i) Pgp expression in mouse brain is limited during late embryogenesis and the newborn period; (ii) Pgp expression increases markedly with postnatal maturation, and (iii) by D21 brain Pgp protein expression approximates adult levels.

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