Spinal cord injuries often occur at the cervical level above the phrenic motor pools, which innervate the diaphragm. The effects of impaired breathing are a leading cause of death from spinal cord ...injuries, underscoring the importance of developing strategies to restore respiratory activity. Here we show that, after cervical spinal cord injury, the expression of chondroitin sulphate proteoglycans (CSPGs) associated with the perineuronal net (PNN) is upregulated around the phrenic motor neurons. Digestion of these potently inhibitory extracellular matrix molecules with chondroitinase ABC (denoted ChABC) could, by itself, promote the plasticity of tracts that were spared and restore limited activity to the paralysed diaphragm. However, when combined with a peripheral nerve autograft, ChABC treatment resulted in lengthy regeneration of serotonin-containing axons and other bulbospinal fibres and remarkable recovery of diaphragmatic function. After recovery and initial transection of the graft bridge, there was an unusual, overall increase in tonic electromyographic activity of the diaphragm, suggesting that considerable remodelling of the spinal cord circuitry occurs after regeneration. This increase was followed by complete elimination of the restored activity, proving that regeneration is crucial for the return of function. Overall, these experiments present a way to markedly restore the function of a single muscle after debilitating trauma to the central nervous system, through both promoting the plasticity of spared tracts and regenerating essential pathways.
Mycobacterial cell wall inhibitors interfere with targets involved in synthesis of mycolic acids, arabinogalactan and peptidoglycan. These antibiotics corrupt structural integrity of the cell ...envelope and this is believed to be the cause of drug mediated cell death. Here, we show that treatment of
BCG with these mechanistically different classes of cell wall inhibitors at MIC caused a 4 to 5-fold increase in intrabacterial ATP concentration. This effect on ATP homeostasis was specific to inhibitors of cell wall synthesis and not observed for other anti-tuberculosis drugs. Treating
BCG with sub-MIC concentrations of the ATP synthase inhibitor bedaquiline or the uncoupler carbonyl cyanide 3-chlorophenylhydrazone suppressed drug induced ATP surge, suggesting that the increase in ATP concentration was due to increased oxidative phosphorylation. Pharmacological suppression of the ATP burst attenuated bactericidal activity of the cell wall-targeting drugs up to 100-fold, suggesting that increased ATP levels are associated with the lethal effect of these antibiotics. Interestingly, inhibition of the ATP burst also suppressed induction of the promoter of the cell envelope stress response operon
by cell wall inhibitors suggesting a link between ATP surge and
expression. In conclusion, we show that treatment of
BCG with inhibitors of cell wall synthesis causes a burst of intrabacterial ATP by increasing oxidative phosphorylation. This ATP surge appears to be required for induction of the
cell envelope stress response operon and to contribute to drug induced cell death. Hence, this work revealed links between inhibition of cell wall synthesis, oxidative phosphorylation,
induction and cell death. The identification of the molecular mechanisms linking these processes may reveal novel targets for the discovery of bactericidal antibiotics.
New drug targets and molecules with bactericidal activity are needed against the respiratory mycobacterial pathogen Mycobacterium abscessus. Employing a lead repurposing strategy, the ...antituberculosis compound GaMF1 was tested against M. abscessus. Whole-cell and ATP synthesis assays demonstrated that GaMF1 inhibits growth and kills M. abscessus by targeting the F-ATP synthase. GaMF1's anti-M. abscessus activity increased in combination with clofazimine, rifabutin, or amikacin. The study expands the repertoire of anti-M. abscessus compounds targeting oxidative phosphorylation.
Unlike Tuberculosis (TB),
lung disease is a highly drug-resistant bacterial infection with no reliable treatment options. De novo
drug discovery is urgently needed but is hampered by the bacterium's ...extreme drug resistance profile, leaving the current drug pipeline underpopulated. One proposed strategy to accelerate de novo
drug discovery is to prioritize screening of advanced TB-active compounds for anti-
activity. This approach would take advantage of the greater chance of homologous drug targets between mycobacterial species, increasing hit rates. Furthermore, the screening of compound series with established structure-activity-relationship, pharmacokinetic, and tolerability properties should fast-track the development of in vitro anti-
hits into lead compounds with in vivo efficacy. In this review, we evaluated the effectiveness of this strategy by examining the literature. We found several examples where the screening of advanced TB chemical matter resulted in the identification of anti-
compounds with in vivo proof-of-concept, effectively populating the
drug pipeline with promising new candidates. These reports validate the screening of advanced TB chemical matter as an effective means of fast-tracking
drug discovery.
Most antibiotics are produced by soil microbes and typically interfere with macromolecular synthesis processes as their antibacterial mechanism of action. These natural products are often large and ...suffer from poor chemical tractability. Here, we discuss discovery, mechanism of action, and the therapeutic potentials of an unusual antibiotic, indole propionic acid (IPA). IPA is produced by the human gut microbiota. The molecule is small, chemically tractable, and targets amino acid biosynthesis. IPA is active against a broad spectrum of mycobacteria, including drug resistant
Mycobacterium tuberculosis
and non-tuberculous mycobacteria (NTM). Interestingly, the microbiota-produced metabolite is detectable in the serum of healthy individuals, tuberculosis (TB) patients, and several animal models. Thus, the microbiota in our gut may influence susceptibility to mycobacterial diseases. If a gut-lung microbiome axis can be demonstrated, IPA may have potential as a biomarker of disease progression, and development of microbiota-based therapies could be explored. In addition to its antimycobacterial activity, the molecule displays anti-inflammatory and antioxidant properties. This raises the possibility that IPA has therapeutic potential as both antibiotic and add-on host-directed drug for the treatment of TB in patient populations where disease morbidity and mortality is driven by excessive inflammation and tissue damage, such as TB-associated immune reconstitution inflammatory syndrome, TB-meningitis, and TB-diabetes.
There exists an abundance of barriers that hinder functional recovery following spinal cord injury, especially at chronic stages. Here, we examine the rescue of breathing up to 1.5 years following ...cervical hemisection in the rat. In spite of complete hemidiaphragm paralysis, a single injection of chondroitinase ABC in the phrenic motor pool restored robust and persistent diaphragm function while improving neuromuscular junction anatomy. This treatment strategy was more effective when applied chronically than when assessed acutely after injury. The addition of intermittent hypoxia conditioning further strengthened the ventilatory response. However, in a sub-population of animals, this combination treatment caused excess serotonergic (5HT) axon sprouting leading to aberrant tonic activity in the diaphragm that could be mitigated via 5HT2 receptor blockade. Through unmasking of the continuing neuroplasticity that develops after injury, our treatment strategy ensured rapid and robust patterned respiratory recovery after a near lifetime of paralysis.
Key points
The functional neuroanatomy of the mammalian respiratory network is far from being understood since experimental tools that measure neural activity across this brainstem‐wide circuit are ...lacking.
Here, we use silicon multi‐electrode arrays to record respiratory local field potentials (rLFPs) from 196–364 electrode sites within 8–10 mm3 of brainstem tissue in single arterially perfused brainstem preparations with respect to the ongoing respiratory motor pattern of inspiration (I), post‐inspiration (PI) and late‐expiration (E2).
rLFPs peaked specifically at the three respiratory phase transitions, E2–I, I–PI and PI–E2.
We show, for the first time, that only the I–PI transition engages a brainstem‐wide network, and that rLFPs during the PI–E2 transition identify a hitherto unknown role for the dorsal respiratory group.
Volumetric mapping of pontomedullary rLFPs in single preparations could become a reliable tool for assessing the functional neuroanatomy of the respiratory network in health and disease.
While it is widely accepted that inspiratory rhythm generation depends on the pre‐Bötzinger complex, the functional neuroanatomy of the neural circuits that generate expiration is debated. We hypothesized that the compartmental organization of the brainstem respiratory network is sufficient to generate macroscopic local field potentials (LFPs), and if so, respiratory (r) LFPs could be used to map the functional neuroanatomy of the respiratory network. We developed an approach using silicon multi‐electrode arrays to record spontaneous LFPs from hundreds of electrode sites in a volume of brainstem tissue while monitoring the respiratory motor pattern on phrenic and vagal nerves in the perfused brainstem preparation. Our results revealed the expression of rLFPs across the pontomedullary brainstem. rLFPs occurred specifically at the three transitions between respiratory phases: (1) from late expiration (E2) to inspiration (I), (2) from I to post‐inspiration (PI), and (3) from PI to E2. Thus, respiratory network activity was maximal at respiratory phase transitions. Spatially, the E2–I, and PI–E2 transitions were anatomically localized to the ventral and dorsal respiratory groups, respectively. In contrast, our data show, for the first time, that the generation of controlled expiration during the post‐inspiratory phase engages a distributed neuronal population within ventral, dorsal and pontine network compartments. A group‐wise independent component analysis demonstrated that all preparations exhibited rLFPs with a similar temporal structure and thus share a similar functional neuroanatomy. Thus, volumetric mapping of rLFPs could allow for the physiological assessment of global respiratory network organization in health and disease.
Key points
The functional neuroanatomy of the mammalian respiratory network is far from being understood since experimental tools that measure neural activity across this brainstem‐wide circuit are lacking.
Here, we use silicon multi‐electrode arrays to record respiratory local field potentials (rLFPs) from 196–364 electrode sites within 8–10 mm3 of brainstem tissue in single arterially perfused brainstem preparations with respect to the ongoing respiratory motor pattern of inspiration (I), post‐inspiration (PI) and late‐expiration (E2).
rLFPs peaked specifically at the three respiratory phase transitions, E2–I, I–PI and PI–E2.
We show, for the first time, that only the I–PI transition engages a brainstem‐wide network, and that rLFPs during the PI–E2 transition identify a hitherto unknown role for the dorsal respiratory group.
Volumetric mapping of pontomedullary rLFPs in single preparations could become a reliable tool for assessing the functional neuroanatomy of the respiratory network in health and disease.
Abstract The hypothalamic paraventricular nucleus (PVN) coordinates autonomic and neuroendocrine systems to maintain homeostasis and to respond to stress. Neuroanatomic and neurophysiologic ...experiments have provided insight into the mechanisms by which the PVN acts. The PVN projects directly to the spinal cord and brainstem and, specifically, to sites that control cardio-respiratory function: the intermediolateral cell columns and phrenic motor nuclei in the spinal cord and rostral ventrolateral medulla (RVLM) and the rostral nuclei in the ventral respiratory column (rVRC) in the brainstem. Activation of the PVN increases ventilation (both tidal volume and frequency) and blood pressure (both heart rate and sympathetic nerve activity). Excitatory and inhibitory neurotransmitters including glutamate and GABA converge in the PVN to influence its neuronal activity. However, a tonic GABAergic input to the PVN directly modulates excitatory outflow from the PVN. Further, even within the PVN, microinjection of GABAA receptor blockers increases glutamate release suggesting an indirect mechanism by which GABA control contributes to PVN functions. PVN activity alters blood pressure and ventilation during various stresses, such as maternal separation, chronic intermittent hypoxia (CIH), dehydration and hemorrhage. Among the several PVN neurotransmitters and neurohormones, vasopressin and oxytocin modulate ventilation and blood pressure. Here, we review our data indicating that increases in vasopressin and vasopressin type 1A (V1A ) receptor signalling in the RVLM and rVRC are mechanisms increasing blood pressure and ventilation after exposure to CIH. That blockade of V1A receptors in the medulla normalizes baseline blood pressure as well as blunts PVN-evoked blood pressure and ventilatory responses in CIH-conditioned animals indicate the role of vasopressin in cardiorespiratory control. In summary, morphological and functional studies have found that the PVN integrates sensory input and projects to the sympathetic and respiratory control systems with descending projections to the medulla and spinal cord.
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