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Effects of [alpha]-bungarotoxin treatment on murine neuromuscular transmission

University of British Columbia

The development and growth of the mammalian neuromuscular junction (NMJ) is orchestrated by complex antero- and retrograde signals exchanged between the nerve terminal and the muscle. The adult NMJ has been shown to display morphological plasticity manifest under conditions of increased or decreased muscle activity, denervation (surgical and pharmacologic) and aging, as part of a continuum of responses to stresses imposed on neuromuscular transmission. Therefore, it seemed plausible to hypothesize that a reduction in postsynaptic sensitivity to ACh with consequent reduced muscle fiber activity would elicit functional changes (an increase or decrease in ACh release) mediated by anterograde and muscle-derived retrograde factors. Using mice, I produced a chronic reduction of functional AChRs by a 3-week, subparalytic treatment with the irreversible AChR antagonist, ot-bungarotoxin. Dose of toxin was adjusted based on the weight and strength of the animals. Strength was assessed by placing the mouse on a rotating rod and monitoring the time it was capable of sustaining itself without falling. Treated animals exhibited a marked reduction in their ability to stay 'roto-rod'. Subsequent electrophysiological experiments were performed on the isolated phrenic nerve/hemidiaphragm preparations to record both spontaneous and evoked ACh release under a variety of Ca and K concentrations and stimulation protocols (30 s 10 Hz period followed by 60 trains-of-ten at 70 Hz at each junction); The chronic treatment with a-bungarotoxin caused an increase in spontaneous and evoked ACh release detected by intracellular microelectrode recordings from the muscle fiber cell. However, the log normal distribution of release was unaltered and neither was its average facilitation in response to train stimulation. Train-evoked release relative to antecedent 10 Hz quantal content or by mean train quantal content was the same in both animal groups but of greater variability in controls. The variance associated with release was found to be similar in both control and treated animals. In summary, I have provided evidence that a chronic reduction in muscle sensitivity to released ACh results in the upregulation of both spontaneous and evoked release, perhaps via the central (CNS) summation and processing of retrograde signals from individual junctions. Our data also suggest that spontaneous ACh release may have a physiological role at the adult NMJ, one of sensing the integrity and level of function of the synapse.

Thesis/Dissertation

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2009-03-12

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http://hdl.handle.net/2429/5961

Pharmacology and Therapeutics

University of British Columbia

UBCV

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