Avaliação comportamental e morfofuncional de junções neuromusculares e músculos estriados esqueléticos de camundongos com disfunção colinérgica a longo prazo

Abstract

The Neuromuscular Junctions (NMJ) is essential for the neuronal communication between the nerve terminals and muscle fibers leading to muscle contraction. One of the most critical processes of cholinergic transmission involves the event of storage of acetylcholine (ACh) in synaptic vesicles (SVs) through the vesicular acetylcholine transporter (VAChT). To assess the functional importance of the VAChT in the cholinergic transmission, Prado et al. (2006) generated a strain of mice knockdown for the VAChT gene and these animals have a pronounced deficit of muscle strength and motor performance. The VAChT knockdown (VAChT KDHOM) mouse model exhibited a ~ 70% decrease in VAChT expression reaches adulthood, but they are myasthenic and present cognitive deficits. In addition, VAChT KDHOM mice present others alterations at the neuromuscular synaptic level such as a reduction in quantal content and size, reduced miniature end-plate potentials frequency, impairment of motor performance and severe deficit in muscle strength. Although some alterations seen in VAChT KDHOM adult mice are already known, there is a lack of information when it comes to cholinergic dysfunction in middle aged and senescence mice. Could it induce further structural and functional changes on pre and postsynaptic components? Thereby, in this work our main goal was to investigate possible pre and postsynaptic alterations in NMJ of the diaphragm muscle and soleus (SOL) and extensor digitorum longus (EDL) muscles from mice with long-term cholinergic deficit. We show that diaphragm from mouse with reduced expression of VAChT does not present major changes in NMJ morphology or pulmonary function but presents alterations in the release of spontaneous synaptic vesicles, amplitude of the miniature end plate potential (MEPP). In addition, we observed that VAChT KDHOM mice performed worse in behavioral motor tests compared to WT littermates with the same age. Accordingly, we found that the SOL muscle fibers from VAChT KDHOM mice presented a significant increase in the cross sectional area (CSA) and expression of the type I Myosin Heavy Chain isoform (MyHC) and EDL presented a reduction in CSA in ours fibers. In addition, we also observed the presence of intramyofibrillar e intermyofibrillar glycogen in SOL muscles from VAChT KDHOM mice that were not found in their WT counterparts. Noteworthy, VAChT KDHOM mice treated with acetylcholinesterase inhibitor pyridostigmine performed similar to WT in motor tests and the CSA of SOL and EDL muscle fibers were partially rescued to control values, suggesting that an increase in ACh availability at the synaptic cleft helps reversing these motor deficits. These data suggest that ACh release plays a distinct role in maintaining the structure of muscular fibers and muscular function general, as well as being important in the regulation of spontaneous release of vesicles.