Piperine activates the thick filament of resting rat skeletal muscle, enhancing dynamic contractility in a fiber-type-dependent manner

Abstract

The myosin-containing thick filament has recently been shown to alter its resting activation level in response to multiple diseases and therapeutics. Changes in thick filament resting activation level are caused by myosin heads transitioning between OFF and ON conformational states. Functionally, this modulation of thick filament activation level is a key regulatory step in muscle contraction and a promising therapeutic target. The availability of resting ON-state myosin heads governs dynamic contractility, which is critical to physical function and well-being. At present, there is a lack of compounds favouring this ON-state in resting skeletal muscle. Piperine is a molecule known to bind to myosin and increase submaximal isometric contractility in fast and slow skeletal muscle. Yet, effects on dynamic contractility and the underlying mechanism responsible for the observed effects in skeletal muscles remain unclear. Here, we used fibre small-angle X-ray diffraction and intact-muscle ex vivo contractility experiments to determine the effects of piperine on resting myosin structure and dynamic contractility in fast and slow rat muscles. X-ray diffraction data suggest that piperine promotes an OFF-to-ON transition of myosin in resting skeletal muscle, increasing the availability of myosin heads for force generation. Functionally, piperine substantially enhanced dynamic contractility in both muscle types, with greater improvements in slow muscle during maximal activation. These findings establish piperine as a tool to probe thick-filament activation in skeletal muscle, highlighting fibre-type-specific effects of thick-filament activation on the recruitment of the contractile reserve capacity. Key Points- Piperine is a compound known to bind to skeletal muscle myosin and enhance isometric contractility in fast and slow muscles, but the underlying molecular mechanisms and effects on dynamic contractile function remain unknown. - We show that piperine increases the activation level of the myosin-containing thick filament in resting fast and slow skeletal muscle, which may explain the effect of piperine on contractile function. - Piperine substantially increases the maximal contractile power of both fast and slow skeletal muscles at low-frequency activation; however, it only enhances the maximal power in slow skeletal muscle at high-frequency activation. - Our data reveal potentiation of dynamic contractility with fibre-type-dependent magnitudes in response to piperine-induced activation of the resting thick filament, a phenomenon that requires further investigation and may ultimately be exploited in the treatment of diseases characterised by muscle weakness. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/689918v2_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1ff3672org.highwire.dtl.DTLVardef@4f809forg.highwire.dtl.DTLVardef@1857404org.highwire.dtl.DTLVardef@83d958_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO Abstract figure legend: We investigated the effects of piperine on 1) the activation level of the resting thick filament and 2) dynamic contractility in fibres and intact slow (soleus) and fast (extensor digitorum longus, EDL) rat muscles, respectively. The activation level of the resting thick filament was assessed pre- and post-piperine incubation using small-angle X-ray diffraction. Dynamic contractility was assessed at submaximal and maximal activation levels by constructing low- and high-frequency force-velocity curves and corresponding power curves using an ex vivo contraction setup. The setup allows for simultaneous experiments on the effects of piperine and vehicle treatment in contralateral muscles. We found that piperine increased the activation level of the resting thick filament by favouring the ON-myosin state in fibres from both muscle types. In whole muscle preparations, piperine also induced substantial increases in dynamic contractility, especially in slow soleus muscle. C_FIG