A chloride channel blocker prevents the suppression by inorganic phosphate of the cytosolic calcium signals that control muscle contraction.

KEY POINTS: Accumulation of inorganic phosphate (Pi ) may contribute to muscle fatigue by precipitating calcium salts inside the sarcoplasmic reticulum (SR). Neither direct demonstration of this process nor definition of the entry pathway of Pi into SR are fully established.  We showed that Pi promoted Ca2+ release at concentrations below 10 mm and decreased it at higher concentrations. This decrease correlated well with that of [Ca2+ ]SR .  Pre-treatment of permeabilized myofibres with 2 mm Cl- channel blocker 9-anthracenecarboxylic acid (9AC) inhibited both effects of Pi .  The biphasic dependence of Ca2+ release on [Pi ] is explained by a direct effect of Pi acting on the SR Ca2+ release channel, combined with the intra-SR precipitation of Ca2+ salts. The effects of 9AC demonstrate that Pi enters the SR via a Cl- pathway of an as-yet-undefined molecular nature. ABSTRACT: Fatiguing exercise causes hydrolysis of phosphocreatine, increasing the intracellular concentration of inorganic phosphate (Pi ). Pi diffuses into the sarcoplasmic reticulum (SR) where it is believed to form insoluble Ca2+ salts, thus contributing to the impairment of Ca2+ release. Information on the Pi entrance pathway is still lacking. In amphibian muscles endowed with isoform 3 of the RyR channel, Ca2+ spark frequency is correlated with the Ca2+ load of the SR and can be used to monitor this variable. We studied the effects of Pi on Ca2+ sparks in permeabilized fibres of the frog. Relative event frequency (f/fref ) rose with increasing [Pi ], reaching 2.54 ± 1.6 at 5 mm, and then decreased monotonically, reaching 0.09 ± 0.03 at [Pi ] = 80 mm. Measurement of [Ca2+ ]SR confirmed a decrease correlated with spark frequency at high [Pi ]. A large [Ca2+ ]SR surge was observed upon Pi removal. Anion channels are a putative path for Pi into the SR. We tested the effect of the chloride channel blocker 9-anthracenecarboxylic acid (9AC) on Pi entrance. 9AC (400 µm) applied to the cytoplasm produced a non-significant increase in spark frequency and reduced the Pi effects on this parameter. Fibre treatment with 2 mm 9AC in the presence of high cytoplasmic Mg2+ suppressed the effects of Pi on [Ca2+ ]SR and spark frequency up to 55 mm [Pi ]. These results suggest that chloride channels (or transporters) provide the main pathway of inorganic phosphate into the SR and confirm that Pi impairs Ca2+ release by accumulating and precipitating with Ca2+ inside the SR, thus contributing to myogenic fatigue.

Muscle synergies during the walk-run and run-walk transitions.

Background: Muscular synergies could represent the patterns of muscular activation used by the central nervous system (CNS) to simplify the production of movement. Studies in walking-running transitions described up to nine synergy modules, and an earlier activation of flexor and extension ankle muscular groups compared to running or walking. Our project aims to study the behaviour of muscle synergies in different stance and swing variations of walking-running (WRT) and running-walking (RWT) transitions. Methods: Twenty-four trained men participated in this study. A variable speed protocol on a treadmill was developed to record the activity of 14 muscle during walking, running and relative transitions. The protocol was based on five ramps of 50 seconds each around ± 10 and 20% of the WRT speed. WRT and RWT were identified according to an abrupt change of the duty factor. Analysing surface electromyography using non-negative matrix factorization (NMF) we obtained synergy modules and temporal activation profiles. Alpha threshold for statistical tests set at 0.05. Results: We described four different transition strides, two for increasing speed transitions, and two for decreasing speed transitions. Four to six synergy modules were found in each condition. According to the maximum cosine similarity results, the two identified WRT conditions shared five modules, while the two RWT conditions shared four modules. WRT and RWT overall shared 4.33 ± 0.58 modules. The activation profiles and centres of activation revealed differences among conditions. Discussion: Transition occurred at step level, and transition strides were composed by walk-like and run-like steps. Compared with previous studies in running and walking, both transitions needed earlier activation of a comparable number of synergy modules. Synergies were affected by acceleration: during RWT the need to dissipate energy, to decrease the speed, was achieved by increasing the number of co-activating muscles. This was reflected in fewer synergy modules and different activation profiles compared to WRT. We believe that our results could be enforced in different applied fields, like clinical gait analysis, physiotherapy and rehabilitation, where plans including co-activation of specific muscular groups could be useful. Gait transitions are common in different sports, and therefore also application in training and sport science would be possible.

Common motor patterns of asymmetrical and symmetrical bipedal gaits.

BACKGROUND: Synergy modules have been used to describe activation of lower limb muscles during locomotion and hence to understand how the system controls movement. Walking and running have been shown shared synergy patterns suggesting common motor control of both symmetrical gaits. Unilateral skipping, an equivalent gait to the quadrupedal gallop in humans, has been defined as the third locomotion paradigm but the use by humans is limited due to its high metabolic cost. Synergies in skipping have been little investigated. In particular, to the best of our knowledge, the joint study of both trailing and leading limbs has never been addressed before. RESEARCH QUESTION: How are organized muscle activation patterns in unilateral skipping? Are they organized in the same way that in symmetrical gaits? If yes, which are the muscle activation patterns in skipping that make it a different gait to walking or running? In the present research, we investigate if there are shared control strategies for all gaits in locomotion. Addressing these questions in terms of muscle synergies could suggest possible determinants of the scarce use of unilateral skipping in humans. METHODS: Electromyographic data of fourteen bilateral muscles were collected from volunteers while performing walking, running and unilateral skipping on a treadmill. Also, spatiotemporal gait parameters were computed from 3D kinematics. The modular composition and activation timing extracted by non-negative matrix factorization were analyzed to detect similarities and differences among symmetrical gaits and unilateral skipping. RESULTS: Synergy modules showed high similarity throughout the different gaits and between trailing and leading limbs during unilateral skipping. The synergy associated with the propulsion force operated by calf muscles was anticipated in bouncing gaits. Temporal features of synergies in the leading leg were very similar to those observed for running. The different role of trailing and leading legs in unilateral skipping was reflected by the different timing in two modules. Activation for weight acceptance was anticipated and extended in the trailing leg, preparing the body for landing impact after the flight phase. A different behaviour was detected in the leading leg, which only deals with a pendular weight transference. SIGNIFICANCE: The evidence gathered in this work supports the hypothesis of shared modules among symmetrical and asymmetrical gaits, suggesting a common motor control despite of the infrequent use of unilateral skipping in humans. Unilateral skipping results from phase-shifted activation of similar muscular groups used in symmetrical gaits, without the need for new muscular groups. The high and anticipated muscle activation in the trailing leg for landing could be the key distinctive event of unilateral skipping.

The voltage sensor of excitation-contraction coupling in mammals: Inactivation and interaction with Ca2.

In skeletal muscle, the four-helix voltage-sensing modules (VSMs) of CaV1.1 calcium channels simultaneously gate two Ca2+ pathways: the CaV1.1 pore itself and the RyR1 calcium release channel in the sarcoplasmic reticulum. Here, to gain insight into the mechanism by which VSMs gate RyR1, we quantify intramembrane charge movement associated with VSM activation (sensing current) and gated Ca2+ release flux in single muscle cells of mice and rats. As found for most four-helix VSMs, upon sustained depolarization, rodent VSMs lose the ability to activate Ca2+ release channels opening; their properties change from a functionally capable mode, in which the mobile sensor charge is called charge 1, to an inactivated mode, charge 2, with a voltage dependence shifted toward more negative voltages. We find that charge 2 is promoted and Ca2+ release inactivated when resting, well-polarized muscle cells are exposed to low extracellular [Ca2+] and that the opposite occurs in high [Ca2+]. It follows that murine VSMs are partly inactivated at rest, which establishes the reduced availability of voltage sensing as a pathogenic mechanism in disorders of calcemia. We additionally find that the degree of resting inactivation is significantly different in two mouse strains, which underscores the variability of voltage sensor properties and their vulnerability to environmental conditions. Our studies reveal that the resting and activated states of VSMs are equally favored by extracellular Ca2+ Promotion by an extracellular species of two states of the VSM that differ in the conformation of the activation gate requires the existence of a second gate, inactivation, topologically extracellular and therefore accessible from outside regardless of the activation state.