Neuromuscular recovery from botulism involves multiple forms of compensatory plasticity

IntroductionBotulinum neurotoxin (BoNT) causes neuroparalytic disease and death by blocking neuromuscular transmission. There are no specific therapies for clinical botulism and the only treatment option is supportive care until neuromuscular function spontaneously recovers, which can take weeks or months after exposure. The highly specialized neuromuscular junction (NMJ) between phrenic motor neurons and diaphragm muscle fibers is the main clinical target of BoNT. Due to the difficulty in eliciting respiratory paralysis without a high mortality rate, few studies have characterized the neurophysiological mechanisms involved in diaphragm recovery from intoxication. Here, we develop a mouse model of botulism that involves partial paralysis of respiratory muscles with low mortality rates, allowing for longitudinal analysis of recovery.Methods and resultsMice challenged by systemic administration of 0.7 LD50 BoNT/A developed physiological signs of botulism, such as respiratory depression and reduced voluntary running activity, that persisted for an average of 8–12 d. Studies in isolated hemidiaphragm preparations from intoxicated mice revealed profound reductions in nerve-elicited, tetanic and twitch muscle contraction strengths that recovered to baseline 21 d after intoxication. Despite apparent functional recovery, neurophysiological parameters remained depressed for 28 d, including end plate potential (EPP) amplitude, EPP success rate, quantal content (QC), and miniature EPP (mEPP) frequency. However, QC recovered more quickly than mEPP frequency, which could explain the discrepancy between muscle function studies and neurophysiological recordings. Hypothesizing that differential modulation of voltage-gated calcium channels (VGCC) contributed to the uncoupling of QC from mEPP frequency, pharmacological inhibition studies were used to study the contributions of different VGCCs to neurophysiological function. We found that N-type VGCC and P/Q-type VGCC partially restored QC but not mEPP frequency during recovery from paralysis, potentially explaining the accelerated recovery of evoked release versus spontaneous release. We identified additional changes that presumably compensate for reduced acetylcholine release during recovery, including increased depolarization of muscle fiber resting membrane potential and increased quantal size.DiscussionIn addition to identifying multiple forms of compensatory plasticity that occur in response to reduced NMJ function, it is expected that insights into the molecular mechanisms involved in recovery from neuromuscular paralysis will support new host-targeted treatments for multiple neuromuscular diseases

The Intranasal Delivery of DNSP-11 and its Effects in Animal Models of Parkinson\u27s Disease

A major challenge in developing disease altering therapeutics for the treatment of Parkinson’s disease (PD) has been the delivery of compounds across the blood-brain barrier (BBB) to the central nervous system (CNS). While direct surgical infusion has been utilized to deliver compounds to the brain that don’t cross the BBB, issues of poor biodistribution in the CNS due in part to properties of the molecules being delivered and/or infusion device protocols have limited the widespread success of this invasive approach. To avoid the issues of surgically delivering compounds to the CNS, numerous studies have examined the use of intranasal administration as a non-invasive delivery method. The data presented in this dissertation examines intranasal administration of dopamine neuron stimulating peptide-11 (DNSP-11), a small, amidated peptide with neuroprotective and restorative properties, and its effects on the nigrostriatal system in animal models of PD. Here we demonstrate that severely lesioned 6-hydroxydopamine (6-OHDA) F344 rats repeatedly administered DNSP-11 intranasally exhibited a decrease in damphetamine- induced rotation, dopamine (DA) turnover, and an increase in tyrosine hydroxylase positive neuronal sparing. Additionally, tracer studies indicated rapid distributed throughout the CNS and CSF following a one-time bilateral intranasal dose of 125I-labeled DNSP-11. These results demonstrate that DNSP-11 can be delivered to the CNS intranasally, and maintains its neuroactive properties on the nigrostriatal system in a rat model of PD. In a dose escalation study of DNSP-11, we evaluated the efficacy of repeated intranasal administration in awake, vertically chaired trained, 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP) hemiparkinsonian rhesus macaques using an atomizer system over a 10-week period. Here we report that animals did not exhibit observable adverse effects at the DNSP-11 concentrations examined, bilateral increases in fine motor performance of the upper limbs, and changes in tissue levels of DA and its metabolites. Finally, tracer studies indicated signal present throughout the CNS and CSF following a one-time bilateral intranasal dose of 125I-labeled DNSP-11. These studies support the efficacy of the repeated intranasal administration of DNSP-11 in awake Rhesus macaques over 10-weeks, while also enhancing motor performance and striatal neurochemistry in a non-human primate model of PD