Florence Picquet

Physiologically adaptive changes of the L5 afferent neurogram and of the rat soleus EMG activity during 14 days of hindlimb unloading and recovery

SUMMARY The hindlimb unloading rat model (HU, Moreys model) is usually used to mimic and study neuromuscular changes that develop during spaceflights. This Earth-based model of microgravity induces a muscular atrophy of the slow postural muscle of hindlimbs, such as the soleus, a loss of strength, modifications of contraction kinetics, changes in histochemical and electrophoretical profiles and modifications of the tonic EMG activity. It has been suggested in the literature that some of these neuromuscular effects were due to a reduction of afferent feedback during HU. However, no direct data have confirmed this hypothesis. The aim of this study was to clearly establish if changes of the L5 afferent neurogram are closely related to the soleus EMG activity during and after 14 days of HU. Immediately after HU, the EMG activity of the soleus muscle disappeared and was associated with a decrease in the afferent neurogram. The soleus electromyographic and afferent activities remained lower than the pre-suspension levels until the sixth day of HU and were recovered between the sixth and the ninth day. On the twelfth and fourteenth days, they were increased beyond the pre-suspension levels. During the first recovery day, these activities were significantly higher than those on the fourteenth HU day and returned to the pre-suspension levels between the third and sixth recovery days. To conclude, our study directly demonstrates that the HU conditions cannot be considered as a functional deafferentation, as suggested in the literature, but only as a reduction of afferent information at the beginning of the HU period.

Compared effects of hindlimb unloading versus terrestrial deafferentation on muscular properties of the rat soleus

Hindlimb unloading is known to induce some neuromuscular changes especially in postural muscles such as the soleus. Our goal was to determine the role of proprioceptive inputs on these modifications by comparing soleus muscle properties of rats being either hindlimb unloaded or terrestrial deafferented. Under deep anesthesia, a first group of rats were submitted to a bilateral deafferentation (DEAF group, n = 6) performed by section of the dorsal roots L(3) to L(5) after laminectomy. A second group of rats was submitted to a hindlimb-unloading period (HU group, n = 6). After 14 days, the morphological and contractile properties as well as the content in myosin heavy-chain (MHC) isoforms were studied in the right soleus muscle in HU and DEAF groups. The results were compared to those obtained in control animals (CON group). After HU versus CON group, the soleus muscle was atrophied and presented a decrease in muscle forces in relation with a slow-to-fast transition characterized by a decrease in kinetic contractile parameters and by an overexpression in the fast MHC isoforms. The DEAF soleus muscle showed both a significant muscle atrophy and a loss of forces when compared with CON rats. The comparison between DEAF and HU rats indicated that some modifications occurring after HU are purely of motor origin (i.e., slow to fast transition), whereas muscle atrophy and decrease in muscle force are partly the result of an afferent silent. Our study underlined the importance of afferent input integrity in the maintenance of muscle characteristics.

Activity-dependent regulation of myelin maintenance in the adult rat

Hindlimb unloading (HU) is known to induce changes in the neuromuscular system. However, no data describing the effects of HU on morphological characteristics of peripheral nerve have been reported so far. Therefore, we used soleus and radial nerves obtained from control and rats submitted to 14 days of HU to study the consequences of a decrease (soleus) or an increase (radial) in neural activity on its morphology. The mean number of fibers was not changed after HU. The soleus nerve axon diameter was weakly affected after HU, whereas the myelin thickness was reduced. For the radial nerve, both axon and fiber diameter were increased, and the myelin thickness and internodal distance were higher in HU rats. These results suggest that regulation of myelin maintenance undergoes plastic mechanisms. Neural activity and/or neural pattern might be essential in the maintenance of myelin sheath in adults.

Hypergravity from conception to adult stage: effects on contractile properties and skeletal muscle phenotype

SUMMARY This study examined the effects of an elevation of the gravity factor (hypergravity – 2 g) on the molecular and functional characteristics of rat soleus and plantaris muscles. Long Evans rats were conceived, born and reared (CBR) continuously in hypergravity conditions until the age of 100 days. Whole muscle morphological parameters, Ca2+ activation characteristics from single skinned fibers, troponin (Tn) subunit and myosin heavy (MHC) and light (MLC) chains isoform compositions were examined in CBR and control muscles from age-paired terrestrial rats. Decreases in body and muscle mass in soleus and plantaris muscles were observed and associated, in the soleus, with a decrease in fiber diameter. The specific force of CBR soleus fibers was increased, and correlated with the elevation of Ca2+ affinity. This was accompanied by slow-to-slower TnC and TnI isoform transitions and a rearrangement in TnT fast isoform content. The MHC transformations of the soleus after hypergravity were associated with the up (down)-regulation of the MHCI (MHCIIa) mRNA isoforms. The MLC2 phosphorylation state remained unchanged in the soleus muscle. The results suggested that the gravity factor could interact with rat muscle development and that hypergravity experiments could provide good tools for the study of myofibrillar protein plasticity and their associated pathways of regulation.

Dendritic spine remodeling induced by hindlimb unloading in adult rat sensorimotor cortex

A sensorimotor restriction, for instance in patients confined to bed, induces an impairment in motor function, which could be due to structural and functional reorganization of the sensorimotor cortex. Hindlimb unloading (HU) is a rodent model used to reproduce the chronic weightless bearing and reduction in hindlimb movement. In this study, we determined whether a 14-day period of HU in adult rats leads to dendritic spine plasticity. For this purpose, we visualized a large number of spines on pyramidal neurons located in superficial and deep layers of the cortex within the hindpaw representation area, by means of confocal microscopy. Spines were classified according to their shape, as stubby, thin, mushroom, or filopodium. Spine density was increased (+26%) after HU. The increase concerned mainly filopodium spines (+82%) and mushrooms (+33%), whereas no change was noticed for stubby and thin spines. Spine length was decreased, whatever their shape. Head diameter evolved differently depending on the layer: it was increased in superficial layers and decreased in deeper ones. These results indicate that morphological changes accompany functional reorganization of motor cortex in response to a decrease in sensorimotor function during adulthood.

Effect of the β2-agonist clenbuterol on the locomotor activity of rat submitted to a 14-day period of hypodynamia-hypokinesia

The beta(2)-adrenergic agonist clenbuterol is known for its anabolic action on normal and atrophied muscles. The aim of this work was to evaluate if chronic clenbuterol administration could prevent alterations in the locomotor activity induced by hindlimb suspension. The effects of clenbuterol were evaluated in three studies: muscle morphological characteristics, observation of locomotor movement and electromyographic activity of soleus and gastrocnemius muscles. Rats were divided into four groups: control (CON, morphological study only), hindlimb suspended (HS), clenbuterol administered (CB, 2 mg kg(-1) per day in drinking water), and hindlimb suspended+clenbuterol administered (HSCB). The soleus muscle weight was reduced in the two suspended groups (HS and HSCB) but did not change after clenbuterol treatment. By contrast, the gastrocnemius weight was not affected by suspension but was increased by clenbuterol (CB and HSCB). Some locomotor deficits were always observed in HS rats (unstable gait, ankle hyperextension, ellipsis). Clenbuterol administration did not prevent these perturbations. Cycle duration and soleus burst duration were increased in the three groups. Soleus mean EMG (burst area/duration) was decreased in HS rats, but not in the two other groups. For the gastrocnemius, burst duration was increased in CB rats, decreased in HSCB rats and unchanged in HS ones; mean EMG did not change. In conclusion, clenbuterol cannot be used as a countermeasure to reduce the alteration in locomotor performance. Moreover, our results suggest that this alteration is specifically related to changes in neuronal properties.

Structure-function relationship of soleus muscle fibres from the rhesus monkey

Functional and structural properties of rhesus monkey skinned fibres were studied in order to examine the relationship between calcium/strontium (Ca/Sr) activation characteristics and protein composition. The fibres were classified according to their Ca/Sr affinity into slow (61%) and fast groups (39%). According to the myosin isoform composition, two additional hybrid types were defined. Thus, four profiles were characterized: two corresponding to slow (S) and fast (F) isoforms and two corresponding to a mixed proportion of slow and fast isoforms. They were called hybrid slow (HS) or hybrid fast (HF) based on the predominant myosin isoform. Tension/pCa parameters and maximal shortening velocities were determined. S fibres showed a higher pCa threshold and affinity as well as shallower slopes of their tension/pCa curve than did F fibres. HS and HF fibres exhibited tension/pCa curves which were positioned close to those of S and F fibres, respectively. No significant difference was observed between S and HS fibres or between F and HF fibres. Maximal shortening velocity values were higher for fibres expressing predominantly fast myosin isoforms. We suggest than when both S and F isoforms of myofibrillar proteins are expressed in a muscle fibre, the functional properties are mainly governed by the predominant isoform.

Effects of unilateral and bilateral labyrinthectomy on rat postural muscle properties: the soleus.

The aim of our study was to determine whether the suppression of the vestibular inputs could have effects on the soleus muscle properties similar to the modifications observed after an episode of microgravity. The inner ear lesion was performed by surgical labyrinthectomy. Twenty-nine male Wistar rats were used for this study and were divided into three experimental groups: control (CONT, n=7), unilateral labyrinthectomized (UL, n=14) and bilateral labyrinthectomized (BL, n=8). Mechanical, histochemical and electrophoretic parameters were determined 17 days after the operation. Furthermore, electromyographic (EMG) activity of the soleus muscle was examined at 1 h, 1 day and 17 days. Our results showed that UL and BL groups did not present any sign of muscle atrophy when compared to CONT group. However, the contractile and phenotypical characteristics of UL and BL soleus muscles revealed that the muscle evolved from slow toward a slower type. This transition was correlated with a more tonic EMG activity pattern. To conclude, our data demonstrated that soleus muscle transformations observed after microgravity (muscle atrophy, slow-to-fast transition, phasic EMG activity) were not directly the consequence of a vestibular silence.

Activity-dependent changes in the electrophysiological properties of regular spiking neurons in the sensorimotor cortex of the rat in vitro.

Sensorimotor performance is highly dependent on the level of physical activity. For instance, a period of disuse induces an impairment of motor performance, which is the result of combined muscular, spinal and supraspinal mechanisms. Concerning this latter origin, our hypothesis was that intrinsic properties and input/output coupling of cells within the sensorimotor cortex might participate to the alteration in cortical motor control. The aim of the present study was thus to examine the basic electrophysiological characteristics of cortical cells in control rats and in animals submitted to 14 days of hindlimb unloading, a model of sensorimotor deprivation. Intracellular recordings were obtained in vitro from coronal slices from cortical hindpaw representation area. We have also made an attempt to determine the morphological characteristics as well as the location of the investigated neurons by biocytin labelling. Passive properties of neurons were affected by hindlimb unloading: input resistance and time constant were decreased (-20%), the rheobase was increased (+34%), whereas the resting potential was unchanged. The frequency-current relationships were also modified, the curve being shifted towards right. The size of body area of recorded neurons was unchanged in unloaded rats. Taken together, these data reflect a decrease in excitability of cortical cells in response to a decreased cortical activation.

Expression of Myosin Heavy Chain Isoforms in Rat Soleus Muscle Spindles After 19 Days of Hypergravity

The aim of this study was to determine whether a period of 19 days in hypergravity was long enough to induce changes in the expression of myosin heavy chain (MyHC) isoforms in the muscle spindles. The soleus muscle of 10 male Wistar rats (control: CONT, n=5; hypergravity: HG, n=5) was frozen, cut into serial sections, and labeled with antibodies against MyHCs: I, IIA, IIA + IIX + IIB, slow-tonic, and α-cardiac. Forty CONT and 45 HG spindles were analyzed. The results from HG spindles compared to CONT showed that there was no change in the cross-sectional area of intrafusal fibers. However, along the entire length of B1 fibers, the expression of both MyHC I and α-cardiac was increased significantly, whereas the labeling against MyHC IIA and MyHC slow-tonic was decreased. In B2 fibers, the labeling against MyHC IIA (region A), slow-tonic (region A), and fast myosins (regions A-C) was statistically decreased. In chain fibers, the labeling against both MyHC IIA and fast MyHC was reduced significantly. We conclude that hypergravity has a real impact on the MyHC content in the muscle spindles and induces some inverse changes of those observed in hypogravity for MyHCs I, α-cardiac, and slow-tonic.