Soleus muscle stability in wild hibernating black bears.

Based on studies of fast skeletal muscles, hibernating black and brown bears resist skeletal muscle atrophy during months of reduced physical activity and not feeding. The present study examined atrophy sparing in the slow soleus muscle, known to be highly prone to disuse atrophy in humans and other mammals. We demonstrated histochemically that the black bear soleus is rich in slow fibers, averaging 84.0 ± 6.6%. The percentages of slow fibers in fall (87.3 ± 4.9%) and during hibernation (87.1 ± 5.6%) did not differ ( P = 0.3152) from summer. The average fiber cross-sectional area to body mass ratio (48.6 ± 11.7 µm2/kg) in winter hibernating bears was not significantly different from that of summer (54.1 ± 11.8 µm2/kg, P = 0.4186) and fall (47.0 ± 9.7 µm2/kg, P = 0.9410) animals. The percentage of single hybrid fibers containing both slow and fast myosin heavy chains, detected biochemically, increased from 2.6 ± 3.8% in summer to 24.4 ± 24.4% ( P = 0.0244) during hibernation. The shortening velocities of individual hybrid fibers remained unchanged from that of pure slow and fast fibers, indicating low content of the minority myosins. Slow and fast fibers in winter bears exhibited elevated specific tension (kN/m2; 22%, P = 0.0161 and 11%, P = 0.0404, respectively) and maintained normalized power. The relative stability of fiber type percentage and size, fiber size-to-body mass ratio, myosin heavy chain isoform content, shortening velocity, power output, and elevated specific tension during hibernation validates the ability of the black bear to preserve the biochemical and performance characteristics of the soleus muscle during prolonged hibernation.

Effects of prolonged space flight on human skeletal muscle enzyme and substrate profiles.

Our primary goal was to determine the effects of 6-mo flight on the International Space Station (ISS) on selected anaerobic and aerobic enzymes, and the content of glycogen and lipids in slow and fast fibers of the soleus and gastrocnemius. Following local anesthesia, biopsies were obtained from nine ISS crew members ∼45 days preflight and on landing day (R+0) postflight. We subdivided the crew into those who ran 200 min/wk or more (high treadmill, HT) in-flight from those who ran <100 min/wk (low treadmill, LT). In the LT group, there was a loss of lipid in soleus type I fibers, and muscle glycogen significantly increased in soleus fiber types postflight. Soleus cytochrome oxidase (CO) activity was significantly depressed postflight in the type I fiber. This was attributed to the LT group where CO activity was reduced 59%. Otherwise, there was no change in the crew mean for type I or IIa fiber glycolytic or mitochondrial enzyme activities pre- vs. postflight in either muscle. However, two of the three HT subjects (Subjects E and H) showed significant increases in both ß-hydroxyacyl-CoA dehydrogenase and citrate synthase in the soleus type I fibers, and Subject E, exhibiting the largest increase in soleus oxidative enzymes, was the only subject to show a significant decrease in glycolytic enzyme activity. It is apparent that crew members performing adequate treadmill running can maintain calf muscle enzymes, which suggests that increased fatigue with weightlessness cannot be directly caused by a decline in muscle enzyme capacity.

Association between maternal periapical lesions and brain inflammation in rat pups.

OBJECTIVE: The objective of this study was to determine whether the presence of maternal tooth periapical lesions was associated with foetal brain inflammation in a pregnant rat model. METHODS: Sprague-Dawley rats were divided into two groups: pregnant rats with induced periapical abscesses (E, n=8) and sham-operated control pregnant rats (S, n=8). The pulps of the first and second maxillary right molars had been exposed and the tooth left open to the oral environment for two weeks prior to initiation of the pregnancy. Following delivery of the pups (E, n=99; S, n=101), each pup was decapitated and the brain was removed and immediately frozen in liquid nitrogen. The tissues were solubilized in PBS containing a protease inhibitor, and norepinephrine (NE), IL-6, IL-1-ß, TNF-α, and myelin basic protein (MBP) were determined by ELISA. Group means were compared by factorial analysis of variance, a post hoc Tukey test, and Pearson's correlation test. p<0.05 was used to reject the null hypothesis. RESULTS: E pups were significantly heavier than S pups. Brain tissue concentrations of IL-6, IL-1-ß, and TNF-α were significantly higher and MBP and norepinephrine concentrations significantly lower in E pups than S pups. Concentrations of IL-6, TNF-α and IL-1-ß were significantly correlated between E serum, pup birthweight, and E pup brain tissue. MBP, NE and IL-6 were significantly correlated within the brain tissues of E pups. CONCLUSION: The data suggest that brain inflammation may be associated with maternal periapical inflammation. This association identifies a modifiable risk factor for adverse pregnancy outcomes.

Effects of gender on serum biomarkers of systemic inflammation coincident to experimentally-induced periapical lesions.

OBJECTIVE: The literature suggests that females have less adverse effects to infection than males, due to the protective effects of oestrogen. The purpose of our study is to compare the systemic effects of induced periapical lesions between groups of animals with various serum concentrations of oestrogen. METHODS: To induce periapical inflammation, two molar tooth pulps were exposed in ovariectomized (OVX) and normal female (F) and castrated (Cast-M) and normal male (M) Sprague-Dawley rats (Experimental group, E). Sham-operated control animals from each group were also studied (Control group, C). Twenty-eight days later, serum and maxillas were collected. Serum 17ß-oestradiol, testosterone, MMP-9, IL-18, IL-6, TNF-α, and IL-1ß concentrations were measured by ELISA. Maxillas were cleaned of residual tissue and digital radiographs were made to verify the presence of periapical lesions. Data were compared by factorial ANOVA, post hoc Tukey, and Pearson correlation tests. Groups were considered to be significantly different when p<0.05. RESULTS: The serum concentration of IL-18, TNF-α, IL-1-ß, IL-6 and MMP-9 was greatest in OVX-E animals, compared to all other groups (p<0.001). F-E rats had significantly higher serum concentrations of these cytokines, compared to F-C. The fold difference in serum concentration of the biomarkers (between E and C groups) was significantly greater in females than males, even though males had higher baseline concentrations of all these biomarkers. CONCLUSION: When females are oestrogen-deficient, their systemic response to periapical lesions is significantly greater than males, suggesting that oestrogen is essential in protecting females from the effects of this type of inflammation.

Prolonged space flight-induced alterations in the structure and function of human skeletal muscle fibres.

The primary goal of this study was to determine the effects of prolonged space flight (180 days) on the structure and function of slow and fast fibres in human skeletal muscle. Biopsies were obtained from the gastrocnemius and soleus muscles of nine International Space Station crew members 45 days pre- and on landing day (R+0) post-flight. The main findings were that prolonged weightlessness produced substantial loss of fibre mass, force and power with the hierarchy of the effects being soleus type I > soleus type II > gastrocnemius type I > gastrocnemius type II. Structurally, the quantitatively most important adaptation was fibre atrophy, which averaged 20% in the soleus type I fibres (98 to 79 µm diameter). Atrophy was the main contributor to the loss of peak force (P(0)), which for the soleus type I fibre declined 35% from 0.86 to 0.56 mN. The percentage decrease in fibre diameter was correlated with the initial pre-flight fibre size (r = 0.87), inversely with the amount of treadmill running (r = 0.68), and was associated with an increase in thin filament density (r = 0.92). The latter correlated with reduced maximal velocity (V(0)) (r = 0.51), and is likely to have contributed to the 21 and 18% decline in V(0) in the soleus and gastrocnemius type I fibres. Peak power was depressed in all fibre types with the greatest loss (55%) in the soleus. An obvious conclusion is that the exercise countermeasures employed were incapable of providing the high intensity needed to adequately protect fibre and muscle mass, and that the crew's ability to perform strenuous exercise might be seriously compromised. Our results highlight the need to study new exercise programmes on the ISS that employ high resistance and contractions over a wide range of motion to mimic the range occurring in Earth's 1 g environment.

Comparative gender differences in local and systemic concentrations of pro-inflammatory cytokines in rats with experimental periodontitis.

BACKGROUND AND OBJECTIVE: There have been few studies of gender differences in response to periodontitis. Thus, we compared gender-specific differences in systemic cytokine concentrations in rats with and without ligature-induced periodontitis. MATERIAL AND METHODS: Experimental periodontal disease was initiated in Sprague-Dawley rats by placing a ligature around the crowns of the second right maxillary molar tooth. Sham-operated control groups were also created. Two weeks later, the right and left maxillary quadrants of teeth, liver and serum were collected from all the rats, and uterine horns were collected from the female rats. Liver and uterine samples were ground in phosphate-buffered saline (10 mg of tissue/mL of phosphate-buffered saline + protease inhibitor) containing a protease inhibitor, and cytokine concentrations were determined by enzyme-linked immunosorbent assay. Digital radiographs were made of maxillary quadrants, and the distance from cemento-enamel junction to alveolar crest was measured using image analysis software. Data were compared by factorial analysis of variance and a post-hoc Tukey test. RESULTS: Female rats with ligatures had greater, but not significantly different, alveolar bone loss than males with ligatures. However, they had higher serum concentrations of interleukin-6, tumor necrosis factor-alpha and C-reactive protein, and liver C-reactive protein (p < 0.05). These females also had higher interleukin-6, tumor necrosis factor-alpha and vascular endothelial growth factor concentrations within the uterine horn, compared to female controls (p < 0.05). Male animals with ligatures had lower serum concentrations of C-reactive protein and higher interleukin-6 and tumor necrosis factor-alpha concentrations within serum, compared to male controls (p < 0.05). CONCLUSION: Our study suggests that females with periodontal disease have a greater risk for inflammatory-based systemic diseases than males.

Relationship between the gingival sulcus depth and interleukin-1 isoform concentrations within the adjacent gingival tissue.

BACKGROUND AND OBJECTIVE: While there is substantial information concerning the concentrations of interleukin-1 isoforms within gingival crevicular fluid, there is little information concerning their concentrations within either normal or diseased gingival tissues. Therefore, the aim of this study was to evaluate the relationship between the concentrations of gingival interleukin-1 isoforms and the adjacent sulcular depth. MATERIAL AND METHODS: Interdental gingival papillae were excised and grouped based on adjacent pocket depth and the presence of bleeding on probing. Gingiva adjacent to a sulcus of < or = 3 mm without bleeding on probing were classified as 'normal'; gingiva adjacent to a 3-mm sulcus with bleeding on probing were classified as 'diseased-slight'; gingiva adjacent to a 4-6-mm sulcus featuring bleeding on probing were classified as 'diseased-moderate'; and gingiva adjacent to a sulcus of > 6 mm featuring bleeding on probing were classified as 'diseased-severe'. Tissues were solublized and the concentrations of interleukin-1beta, interleukin-1alpha, interleukin-1 receptor antagonist and interleukin-6 were assessed by enzyme-linked immunosorbent assay. Data were compared by factorial analysis of variance, the post-hoc Tukey test and the Pearson's correlation test. RESULTS: Gingival concentrations of interleukin-6, interleukin-1 receptor antagonist, interleukin-1alpha- and interleukin-1beta were significantly greater at diseased-severe sites than at normal, diseased-slight, or diseased-moderate sites (p < 0.05); the gingival concentrations of interleukin-1 receptor antagonist and interleukin-1alpha were significantly greater at diseased-severe than at diseased-moderate sites (p < 0.05). Interleukin-1 receptor antagonist concentrations were significantly correlated with both interleukin-1alpha and interleukin-1beta concentrations. The ratios of concentrations of the interleukin-1 isoforms were different at the various stages of inflammation. CONCLUSION: Our data indicated a progressive increase in gingival concentrations of interleukin-1 isoforms with increased adjacent sulcular depth. However, within 'diseased' tissues, the proportional concentrations of interleukin-1alpha and -beta to interleukin-1 receptor antagonist were lowest within diseased-severe tissues.

Effects of induced periapical abscesses on rat pregnancy outcomes.

OBJECTIVE: The objective of this study was to determine the effects of induced periapical abscesses on pregnant rats. DESIGN: In 1/2 of the animals (n=16), the pulps of the maxillary right molars were exposed to the oral environment, which resulted in a periapical abscess. The other 1/2 (n=16) were sham-operated. 1/2 of the animals of both groups became pregnant 2 weeks later. The pregnancy duration, and weight and number of pups were assessed at delivery. Serum, liver and uterine horn samples were taken from all animals at euthanasia and serum IL-6, endothelin-1, TNF-alpha, IL-10, cortisol and insulin were determined by ELISA. Liver concentrations of IL-6, CRP and IL-6 and uterine horn concentrations of IL-6, vascular endothelial growth factor (VEGF), TNF-alpha, IL-10 and IL-1-beta were assessed by ELISA. Blood glucose concentrations were determined using a glucometer. Outcome variables were compared by factorial ANOVA, a post hoc Tukey test, and Pearson's correlation test. RESULTS: Pregnant rats with periapical abscesses had a significantly longer pregnancy and delivered pups with a significantly higher birthweight (p<0.05). They had significantly higher concentrations of IL-6, VEGF, IL-1-beta, and IL-10 within the uterine horn and IL-6, CRP and TNF-alpha within the liver (p<0.01). Blood glucose and serum TNF-alpha, IL-6, endothelin-1, IL-10, and insulin concentrations were significantly higher in the pregnant animals with pulpal abscesses (p<0.01). CONCLUSION: The significant increase in serum TNF-alpha, taken together with significant increases in blood glucose and serum insulin concentrations, suggest that animals with induced periapical abscesses developed insulin resistance, which significantly affected their pregnancy outcomes.

Skeletal muscle fiber atrophy: altered thin filament density changes slow fiber force and shortening velocity.

Single skinned fibers from soleus and adductor longus (AL) muscles of weight-bearing control rats and rats after 14-day hindlimb suspension unloading (HSU) were studied physiologically and ultrastructurally to investigate how slow fibers increase shortening velocity (V0) without fast myosin. We hypothesized that unloading and shortening of soleus during HSU reduces densities of thin filaments, generating wider myofilament separations that increase V0 and decrease specific tension (kN/m2). During HSU, plantarflexion shortened soleus working length 23%. AL length was unchanged. Both muscles atrophied as shown by reductions in fiber cross-sectional area. For AL, the 60% atrophy accounted fully for the 58% decrease in absolute tension (mN). In the soleus, the 67% decline in absolute tension resulted from 58% atrophy plus a 17% reduction in specific tension. Soleus fibers exhibited a 25% reduction in thin filaments, whereas there was no change in AL thin filament density. Loss of thin filaments is consistent with reduced cross bridge formation, explaining the fall in specific tension. V0 increased 27% in soleus but was unchanged in AL. The V0 of control and HSU fibers was inversely correlated (R = -0.83) with thin filament density and directly correlated (R = 0.78) with thick-to-thin filament spacing distance in a nonlinear fashion. These data indicate that reduction in thin filament density contributes to an increased V0 in slow fibers. Osmotically compacting myofilaments with 5% dextran returned density, spacing, and specific tension and slowed V0 to near-control levels and provided evidence for myofilament spacing modulating tension and V0.

Passive stretch inhibits central corelike lesion formation in the soleus muscles of hindlimb-suspended unloaded rats.

Hindlimb suspension unloading (HSU) is a ground-based model simulating the effects of microgravity unloading on the musculoskeletal system. In this model, gravity causes the hind foot of the rat to drop, opening the front of the ankle to 90-105 degrees plantar flexion at rest. As HSU proceeds, the normal weight-bearing angle of 30 degrees dorsiflexion is achieved progressively less, and the contraction range of soleus is abbreviated. Our laboratory reported that 12 days of HSU caused central corelike lesions (CCLs) of myofibril breakdown (Riley DA, Slocum GR, Bain JL, Sedlak FR, Sowa TE, and Mellender JW. J Appl Physiol. 69: 58-66, 1990). The present study investigated whether daily stretch of the calf muscles prevents CCL formation. The soleus muscles of HSU Sprague-Dawley male rats (approximately 287 g) were lengthened by unilateral ankle splinting at 30 degrees. Compared with the nonsplinted side, splinting for 10 or 20 min per day in awake rats significantly decreased CCLs in soleus by 88 and 91%, respectively (P < 0.01). Compared with control muscle wet weight, 20-min splinting reduced atrophy by 33%, whereas 10-min splinting ameliorated atrophy by 17% (P < 0.01). Bilateral soleus electromyograph recording revealed higher levels of contractile activity on the splinted side during splinting. To isolate the effects of stretch from isometric contractile activity, contractions were eliminated by whole animal anesthesia with isoflurane during 10-min daily splinting. The percentage of fibers with CCLs was reduced by 57%, and the average lesion size was 29% smaller in the stretched muscle (P < 0.05). Soleus muscle wet weight and fiber area were unaltered by stretch alone. Loaded contractions during splinting are necessary to prevent muscle fiber atrophy. Passive muscle stretch acts to maintain myofibril structural integrity.

Functional properties of slow and fast gastrocnemius muscle fibers after a 17-day spaceflight.

The purpose of this investigation was to study the effects of a 17-day spaceflight on the contractile properties of individual fast- and slow-twitch fibers isolated from biopsies of the fast-twitch gastrocnemius muscle of four male astronauts. Single chemically skinned fibers were studied during maximal Ca2+-activated contractions with fiber myosin heavy chain (MHC) isoform expression subsequently determined by SDS gel electrophoresis. Spaceflight had no significant effect on the mean diameter or specific force of single fibers expressing type I, IIa, or IIa/IIx MHC, although a small reduction in average absolute force (P(o)) was observed for the type I fibers (0.68 +/- 0.02 vs. 0.64 +/- 0.02 mN, P < 0.05). Subject-by-flight interactions indicated significant intersubject variation in response to the flight, as postflight fiber diameter and P(o) where significantly reduced for the type I and IIa fibers obtained from one astronaut and for the type IIa fibers from another astronaut. Average unloaded shortening velocity [V(o), in fiber lengths (FL)/s] was greater after the flight for both type I (0.60 +/- 0.03 vs. 0.76 +/- 0.02 FL/s) and IIa fibers (2.33 +/- 0.25 vs. 3.10 +/- 0.16 FL/s). Postflight peak power of the type I and IIa fibers was significantly reduced only for the astronaut experiencing the greatest fiber atrophy and loss of P(o). These results demonstrate that 1) slow and fast gastrocnemius fibers show little atrophy and loss of P(o) but increased V(o) after a typical 17-day spaceflight, 2) there is, however, considerable intersubject variation in these responses, possibly due to intersubject differences in in-flight physical activity, and 3) in these four astronauts, fiber atrophy and reductions in P(o) were less for slow and fast fibers obtained from the phasic fast-twitch gastrocnemius muscle compared with slow and fast fibers obtained from the slow antigravity soleus [J. J. Widrick, S. K. Knuth, K. M. Norenberg, J. G. Romatowski, J. L. W. Bain, D. A. Riley, M. Karhanek, S. W. Trappe, T. A. Trappe, D. L. Costill, and R. H. Fitts. J Physiol (Lond) 516: 915-930, 1999].

Effects of hindlimb unloading on neuromuscular development of neonatal rats.

We hypothesized that hindlimb suspension unloading of 8-day-old neonatal rats would disrupt the normal development of muscle fiber types and the motor innervation of the antigravity (weightbearing) soleus muscles but not extensor digitorum longus (EDL) muscles. Five rats were suspended 4.5 h and returned 1.5 h to the dam for nursing on a 24 h cycle for 9 days. To control for isolation from the dam, the remaining five littermates were removed on the same schedule but not suspended. Another litter of 10 rats housed in the same room provided a vivarium control. Fibers were typed by myofibrillar ATPase histochemistry and immunostaining for embryonic, slow, fast IIA and fast IIB isomyosins. The percentage of multiple innervation and the complexity of singly-innervated motor terminal endings were assessed in silver/cholinesterase stained sections. Unique to the soleus, unloading accelerated production of fast IIA myosin, delayed expression of slow myosin and retarded increases in standardized muscle weight and fiber size. Loss of multiple innervation was not delayed. However, fewer than normal motor nerve endings achieved complexity. Suspended rats continued unloaded hindlimb movements. These findings suggest that motor neurons resolve multiple innervation through nerve impulse activity, whereas the postsynaptic element (muscle fiber) controls endplate size, which regulates motor terminal arborization. Unexpectedly, in the EDL of unloaded rats, transition from embryonic to fast myosin expression was retarded. Suspension-related foot drop, which stretches and chronically loads EDL, may have prevented fast fiber differentiation. These results demonstrate that neuromuscular development of both weightbearing and non-weightbearing muscles in rats is dependent upon and modulated by hindlimb loading.

Decreased thin filament density and length in human atrophic soleus muscle fibers after spaceflight.

Soleus muscle fibers were examined electron microscopically from pre- and postflight biopsies of four astronauts orbited for 17 days during the Life and Microgravity Sciences Spacelab Mission (June 1996). Myofilament density and spacing were normalized to a 2. 4-microm sarcomere length. Thick filament density ( approximately 1, 062 filaments/microm(2)) and spacing ( approximately 32.5 nm) were unchanged by spaceflight. Preflight thin filament density (2, 976/microm(2)) decreased significantly (P < 0.01) to 2,215/microm(2) in the overlap A band region as a result of a 17% filament loss and a 9% increase in short filaments. Normal fibers had 13% short thin filaments. The 26% decrease in thin filaments is consistent with preliminary findings of a 14% increase in the myosin-to-actin ratio. Lower thin filament density was calculated to increase thick-to-thin filament spacing in vivo from 17 to 23 nm. Decreased density is postulated to promote earlier cross-bridge detachment and faster contraction velocity. Atrophic fibers may be more susceptible to sarcomere reloading damage, because force per thin filament is estimated to increase by 23%.

Effect of a 17 day spaceflight on contractile properties of human soleus muscle fibres.

1. Soleus biopsies were obtained from four male astronauts 45 days before and within 2 h after a 17 day spaceflight. 2. For all astronauts, single chemically skinned post-flight fibres expressing only type I myosin heavy chain (MHC) developed less average peak Ca2+ activated force (Po) during fixed-end contractions (0.78 +/- 0. 02 vs. 0.99 +/- 0.03 mN) and shortened at a greater mean velocity during unloaded contractions (Vo) (0.83 +/- 0.02 vs. 0.64 +/- 0.02 fibre lengths s-1) than pre-flight type I fibres. 3. The flight-induced decline in absolute Po was attributed to reductions in fibre diameter and/or Po per fibre cross-sectional area. Fibres from the astronaut who experienced the greatest relative loss of peak force also displayed a reduction in Ca2+ sensitivity. 4. The elevated Vo of the post-flight slow type I fibres could not be explained by alterations in myosin heavy or light chain composition. One alternative possibility is that the elevated Vo resulted from an increased myofilament lattice spacing. This hypothesis was supported by electron micrographic analysis demonstrating a reduction in thin filament density post-flight. 5. Post-flight fibres shortened at 30 % higher velocities than pre-flight fibres at external loads associated with peak power output. This increase in shortening velocity either reduced (2 astronauts) or prevented (2 astronauts) a post-flight loss in fibre absolute peak power (microN (fibre length) s-1). 6. The changes in soleus fibre diameter and function following spaceflight were similar to those observed after 17 days of bed rest. Although in-flight exercise countermeasures probably reduced the effects of microgravity, the results support the idea that ground-based bed rest can serve as a model of human spaceflight. 7. In conclusion, 17 days of spaceflight decreased force and increased shortening velocity of single Ca2+-activated muscle cells expressing type I MHC. The increase in shortening velocity greatly reduced the impact that impaired force production had on absolute peak power.

Disproportionate loss of thin filaments in human soleus muscle after 17-day bed rest.

Previously we reported that, after 17-day bed rest unloading of 8 humans, soleus slow fibers atrophied and exhibited increased velocity of shortening without fast myosin expression. The present ultrastructural study examined fibers from the same muscle biopsies to determine whether decreased myofilament packing density accounted for the observed speeding. Quantitation was by computer-assisted morphometry of electron micrographs. Filament densities were normalized for sarcomere length, because density depends directly on length. Thick filament density was unchanged by bed rest. Thin filaments/microm2 decreased 16-23%. Glycogen filled the I band sites vacated by filaments. The percentage decrease in thin filaments (Y) correlated significantly (P < 0.05) with the percentage increase in velocity (X), (Y = 0.1X + 20%, R2 = 0.62). An interpretation is that fewer filaments increases thick to thin filament spacing and causes earlier cross-bridge detachment and faster cycling. Increased velocity helps maintain power (force x velocity) as atrophy lowers force. Atrophic muscles may be prone to sarcomere reloading damage because force/microm2 was near normal, and force per thin filament increased an estimated 30%.

Effect of 17 days of bed rest on peak isometric force and unloaded shortening velocity of human soleus fibers.

The purpose of this study was to examine the effect of prolonged bed rest (BR) on the peak isometric force (P0) and unloaded shortening velocity (V0) of single Ca(2+)-activated muscle fibers. Soleus muscle biopsies were obtained from eight adult males before and after 17 days of 6 degrees head-down BR. Chemically permeabilized single fiber segments were mounted between a force transducer and position motor, activated with saturating levels of Ca2+, and subjected to slack length steps. V0 was determined by plotting the time for force redevelopment vs. the slack step distance. Gel electrophoresis revealed that 96% of the pre- and 87% of the post-BR fibers studied expressed only the slow type I myosin heavy chain isoform. Fibers with diameter > 100 microns made up only 14% of this post-BR type I population compared with 33% of the pre-BR type I population. Consequently, the post-BR type I fibers (n = 147) were, on average, 5% smaller in diameter than the pre-BR type I fibers (n = 218) and produced 13% less absolute P0. BR had no overall effect on P0 per fiber cross-sectional area (P0/CSA), even though half of the subjects displayed a decline of 9-12% in P0/CSA after BR. Type I fiber V0 increased by an average of 34% with BR. Although the ratio of myosin light chain 3 to myosin light chain 2 also rose with BR, there was no correlation between this ratio and V0 for either the pre- or post-BR fibers. In separate fibers obtained from the original biopsies, quantitative electron microscopy revealed a 20-24% decrease in thin filament density, with no change in thick filament density. These results raise the possibility that alterations in the geometric relationships between thin and thick filaments may be at least partially responsible for the elevated V0 of the post-BR type I fibers.

In-flight and postflight changes in skeletal muscles of SLS-1 and SLS-2 spaceflown rats.

Spacelab Life Sciences-1 and -2 provided skeletal muscles from rats dissected in flight for the first time and 2 h to 14 days postflight. The muscles permitted the distinguishing of primary adaptations to microgravity from secondary reloading-induced alterations. In microgravity, rats adopted bipedal forelimb locomotion with the hindlimbs relegated to grasping activities. On landing day, body posture was abnormally low and walking was stilted at a rate one-third of normal. The adductor longus (AL) and soleus muscles exhibited decreased myofiber areas that did not recover 14 days postflight. Doubling of the nonmyofiber area indicated interstitial edema in AL muscles 2.3 h postflight. Solei did not manifest edema postflight, and neither muscle showed edema in flight. Sarcomere eccentric contraction-like lesions were detected in 2.6% of AL myofibers 4.5 h postflight; lesions were absent earlier postflight and in flight. At 9 days postflight, these lesions were repaired but regenerating AL myofibers were present, which suggests that myofiber necrosis occurred 1-2 days postflight. These studies demonstrate that muscle atrophy occurs in microgravity, whereas interstitial edema and sarcomere lesions are postflight phenomena.

Transformation of cardiac endothelium into cushion mesenchyme is dependent on ES/130: temporal, spatial, and functional studies in the early chick embryo.

ES/130 is a novel 130-kDa protein that has been linked previously to the transformation of endocardial endothelium into cushion mesenchyme. In the present study we report the localization of protein and mRNA for ES/130 in stages 7-plus through 20 chick embryos and present functional data related to a potential mechanism for ES/130. The temporal and spatial regulation of ES/130 expression suggests that this epithelial-to-mesenchymal transformation is a result of homogenetic induction. Functional studies indicate that myocardially derived ES/130 elicits expression of this protein by target AV endothelial cells, which is linked to a signal transduction cascade. The localization of ES/130 to other sites of inductive interactions (e.g., limb bud ectoderm, gut, and notochord) implies that this protein may have a more widespread importance to embryogenesis beyond its involvement in cardiac cushion tissue formation.

Histochemical carbonic anhydrase in rat inner medullary collecting duct.

Rat inner medullary collecting duct (IMCD) secretes substantial amounts of H+. However, carbonic anhydrase (CA), a concomitant of H+ secretion, has been generally reported absent in this segment. To reexamine this problem, we investigated CA and the morphological phenotypes of cells comprising the IMCD by CA histochemistry, using a modified Hansson technique with light and electron microscopy. Throughout the medulla, tubule cells exhibit histochemical CA activity. In the initial third of the inner medulla, a small proportion have features of intercalated cells and demonstrate some degree of CA activity. However, the majority population in the early portions of the IMCD appears to consist of principal cells. These also show CA staining of widely variable intensity, both among and within cells. A third cell type, previously called "IMCD cells", appears in the middle portion of the IMCD and is the only cell type present near the papilla tip. In contrast to previous reports, these "IMCD cells" have histochemical CA staining, also of highly variable intensity. These results demonstrate that stainable carbonic anhydrase to support acidification is present throughout the rat IMCD, both in intercalated cells and in some cells clearly not of this type. Therefore, the presence of CA is not specific for the intercalated cell type and suggests that other cell types may participate in acid secretion in IMCD.