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Combined retrograde tracing (using fluorescent tracer Fast blue) and double-labelling immunofluorescence were used to study the distribution and immunohistochemical characteristics of neurons projecting to the trapezius muscle in mature male rats (n=9). As revealed by retrograde tracing, Fast blue-positive (FB+) neurons were located within the ambiguous nucleus and accessory nucleus of the grey matter of the spinal cord. Immunohistochemistry revealed that nearly all the neurons were cholinergic in nature [choline acetyltransferase (ChAT)-positive]. Retrogradely labelled neurons displayed also immunoreactivities to calcitonin gene-related peptide (CGRP; approximately 60% of FB+ neurons), nitric oxide synthase (NOS; 50%), substance P (SP; 35%), Leu5-Enkephalin (LEnk; 10%) and vasoactive intestinal polypeptide (VIP; 5%). The analysis of double-stained tissue sections revealed that all CGRP-, VIP- and LEnk-immunoreactive FB+ perikarya were simultaneously ChAT-positive. The vast majority of the neurons expressing SP- or NOS-immunoreactivity were also cholinergic in nature; however, solitary somata were ChAT-negative. FB+ perikarya were surrounded by numerous varicose nerve fibres (often forming basket-like structures) immunoreactive to LEnk or SP. They were also associated with some CGRP-, NOS- and neuropeptide Y-positive nerve terminals.
During natural contractions of a muscle, motor units (MUs) are activated by irregular discharge patterns of motoneurones. The aim of this study was to analyze changes in contractile forces of MUs following patterns of stimulation at variable frequency. Experiments were performed on 33 functionally isolated MUs of the fast-type in the medial gastrocnemius (MG) muscle of adult Wistar rats, under pentobarbital anaesthesia. The MUs forces evoked at five different regular stimulation patterns of constant frequencies were compared to forces generated during five random patterns of irregular stimulation with the same mean values of interpulse intervals, between 10 and 75 ms, and variability of these intervals of ± 50% in each case. These values cover the natural range of the preferred firing rates of the MG motoneurones from unfused to nearly fused tetanic contractions. Analysis of changes in tetanic forces indicated a linear relationship between the interpulse interval as well as the initial level of the force and the amplitude of the force increase of the next contraction. It was demonstrated that variability of the instantaneous tetanic force during the irregular discharge pattern depends on the level of tetanic fusion. Moreover, it was demonstrated that for low and moderately-fused tetani, effectiveness of a MU contraction (expressed as the force-time area) is considerably higher for contractions evoked by irregular stimulation patterns. On the basis of the results of this study it was supposed that during voluntary contractions, the influence of changes in the motoneuronal firing rate on the motor unit force depends on the initial level of force.
Pharmacological up-regulation of heat shock proteins (hsps) rescues motoneurons from cell death in a mouse model of amyotrophic lateral sclerosis. However, the relationship between increased hsp expression and neuronal survival is not straightforward. Here we examined the effects of two pharmacological agents that induce the heat shock response via activation of HSF-1, on stressed primary motoneurons in culture. Although both arimoclomol and celastrol induced the expression of Hsp70, their effects on primary motoneurons in culture were significantly different. Whereas arimoclomol had survival-promoting effects, rescuing motoneurons from staurosporin and H2O2 induced apoptosis, celastrol not only failed to protect stressed motoneurons from apoptosis under same experimental conditions, but was neurotoxic and induced neuronal death. Immunostaining of celastrol-treated cultures for hsp70 and activated caspase-3 revealed that celastrol treatment activates both the heat shock response and the apoptotic cell death cascade. These results indicate that not all agents that activate the heat shock response will necessarily be neuroprotective.
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