Muscle phenotype remains unaltered after limb autotomy and unloading.

Loss of chelipeds in crustaceans results in severe atrophy of the major muscle responsible for lifting the limb, the anterior levator. We decided to test if this loss of mechanical load altered muscle phenotype as measured by SDS-PAGE analysis of levator total protein and actomyosin fractions. Levator muscles of adult crayfish, Procambarus clarkii, with either functional regenerate limbs or lack of limb buds (papilla stage) were compared with those from normal contralateral limbs and those from pristine animals. We find that there is no difference in protein profiles among the three conditions. However, the total protein profile for the dually excited levator muscle is unique compared to those of fast or slow muscles of the abdomen (L and SEL, respectively), which receive only phasic or tonic excitatory innervation. The levator myosin heavy chain profile is similar to that of slow phenotype muscles such as the SEL and opener. We conclude that load does not influence levator phenotype. This is likely due either to the intact innervation and continued activation of the levator during atrophy or to the maintenance of passive tension on the muscle. J. Exp. Zool. 289:10-22, 2001.

Sensitivity of transformed (phasic to tonic) motor neurons to the neuromodulator 5-HT.

Long-term adaptation resulting in a 'tonic-like' state can be induced in phasic motor neurons of the crayfish, Procambarus clarkii, by daily low-frequency stimulation [Lnenicka, G.A., Atwood, H.L., 1985b. Long-term facilitation and long-term adaptation at synapses of a crayfish phasic motoneuron. J. Neurobiol. 16, 97-110]. To test the hypothesis that motor neurons undergoing adaptation show increased responses to the neuromodulator serotonin (5-HT), phasic motor neurons innervating the deep abdominal extensor muscles of crayfish were stimulated at 2.5 Hz, 2 h/day, for 7 days. One day after cessation of conditioning, contralateral control and conditioned motor neurons of the same segment were stimulated at 1 Hz and the induced excitatory post-synaptic potentials (EPSPs) were recorded from DEL(1) muscle fibers innervated by each motor neuron type. Recordings were made in saline without and with 100 nM 5-HT. EPSP amplitudes were increased by 5-HT exposure in all cases. Conditioned muscles exposed to 5-HT showed a 2-fold higher percentage of increase in EPSP amplitude than did control muscles. Thus, the conditioned motor neurons behaved like intrinsically tonic motoneurons in their response to 5-HT. While these results show that long-term adaptation (LTA) extends to 5-HT neuromodulation, no phenotype switch could be detected in the postsynaptic muscle. Protein isoform profiles, including the myosin heavy chains, do not change after 1 week of conditioning their innervating motor neurons.

Muscle type-specific myosin isoforms in crustacean muscles.

Differential expression of multiple myosin heavy chain (MyHC) genes largely determines the diversity of critical physiological, histochemical, and enzymatic properties characteristic of skeletal muscle. Hypotheses to explain myofiber diversity range from intrinsic control of expression based on myoblast lineage to extrinsic control by innervation, hormones, and usage. The unique innervation and specialized function of crayfish (Procambarus clarkii) appendicular and abdominal musculature provide a model to test these hypotheses. The leg opener and superficial abdominal extensor muscles are innervated by tonic excitatory motoneurons. High resolution SDS-PAGE revealed that these two muscles express the same MyHC profile. In contrast, the deep abdominal extensor muscles, innervated by phasic motoneurons, express MyHC profiles different from the tonic profiles. The claw closer muscles are dually innervated by tonic and phasic motoneurons and a mixed phenotype was observed, albeit biased toward the phasic profile seen in the closer muscle. These results indicate that multiple MyHC isoforms are present in the crayfish and that differential expression is associated with diversity of muscle type and function.