Muscle 4EBP1 activation modifies the structure and function of the neuromuscular junction in mice.

Dysregulation of mTOR complex 1 (mTORC1) activity drives neuromuscular junction (NMJ) structural instability during aging; however, downstream targets mediating this effect have not been elucidated. Here, we investigate the roles of two mTORC1 phosphorylation targets for mRNA translation, ribosome protein S6 kinase 1 (S6K1) and eukaryotic translation initiation factor 4E-binding protein 1 (4EBP1), in regulating NMJ structural instability induced by aging and sustained mTORC1 activation. While myofiber-specific deletion of S6k1 has no effect on NMJ structural integrity, 4EBP1 activation in murine muscle induces drastic morphological remodeling of the NMJ with enhancement of synaptic transmission. Mechanistically, structural modification of the NMJ is attributed to increased satellite cell activation and enhanced post-synaptic acetylcholine receptor (AChR) turnover upon 4EBP1 activation. Considering that loss of post-synaptic myonuclei and reduced NMJ turnover are features of aging, targeting 4EBP1 activation could induce NMJ renewal by expanding the pool of post-synaptic myonuclei as an alternative intervention to mitigate sarcopenia.

Sarcopenia: Tilting the Balance of Protein Homeostasis.

Sarcopenia, defined as age-associated decline of muscle mass and function, is a risk factor for mortality and disability, and comorbid with several chronic diseases such as type II diabetes and cardiovascular diseases. Clinical trials showed that nutritional supplements had positive effects on muscle mass, but not on muscle function and strength, demonstrating our limited understanding of the molecular events involved in the ageing muscle. Protein homeostasis, the equilibrium between protein synthesis and degradation, is proposed as the major mechanism underlying the development of sarcopenia. As the key central regulator of protein homeostasis, the mammalian target of rapamycin (mTOR) is proposed to be essential for muscle hypertrophy. Paradoxically, sustained activation of mTOR complex 1 (mTORC1) is associated with a loss of sensitivity to extracellular signaling in the elderly. It is not understood why sustained mTORC1 activity, which should induce muscle hypertrophy, instead results in muscle atrophy. Here, recent findings on the implications of disrupting protein homeostasis on muscle physiology and sarcopenia development in the context of mTOR/protein kinase B (AKT) signaling are reviewed. Understanding the role of these molecular mechanisms during the ageing process will contribute towards the development of targeted therapies that will improve protein metabolism and reduce sarcopenia.

S-Nitrosylation of Divalent Metal Transporter 1 Enhances Iron Uptake to Mediate Loss of Dopaminergic Neurons and Motoric Deficit.

Elevated iron deposition has been reported in Parkinson's disease (PD). However, the route of iron uptake leading to high deposition in the substantia nigra is unresolved. Here, we show a mechanism in enhanced Fe2+ uptake via S-nitrosylation of divalent metal transporter 1 (DMT1). While DMT1 could be S-nitrosylated by exogenous nitric oxide donors, in human PD brains, endogenously S-nitrosylated DMT1 was detected in postmortem substantia nigra. Patch-clamp electrophysiological recordings and iron uptake assays confirmed increased Mn2+ or Fe2+ uptake through S-nitrosylated DMT1. We identified two major S-nitrosylation sites, C23 and C540, by mass spectrometry, and DMT1 C23A or C540A substitutions abolished nitric oxide (NO)-mediated DMT1 current increase. To evaluate in vivo significance, lipopolysaccharide (LPS) was stereotaxically injected into the substantia nigra of female and male mice to induce inflammation and production of NO. The intranigral LPS injection resulted in corresponding increase in Fe2+ deposition, JNK activation, dopaminergic neuronal loss and deficit in motoric activity, and these were rescued by the NO synthase inhibitor l-NAME or by the DMT1-selective blocker ebselen. Lentiviral knockdown of DMT1 abolished LPS-induced dopaminergic neuron loss.SIGNIFICANCE STATEMENT Neuroinflammation and high cytoplasmic Fe2+ levels have been implicated in the initiation and progression of neurodegenerative diseases. Here, we report the unexpected enhancement of the functional activity of transmembrane divalent metal transporter 1 (DMT1) by S-nitrosylation. We demonstrated that S-nitrosylation increased DMT1-mediated Fe2+ uptake, and two cysteines were identified by mass spectrometry to be the sites for S-nitrosylation and for enhanced iron uptake. One conceptual advance is that while DMT1 activity could be increased by external acidification because the gating of the DMT1 transporter is proton motive, we discovered that DMT1 activity could also be enhanced by S-nitrosylation. Significantly, lipopolysaccharide-induced nitric oxide (NO)-mediated neuronal death in the substantia nigra could be ameliorated by using l-NAME, a NO synthase inhibitor, or by ebselen, a DMT1-selective blocker.

The forebrain medial septal region and nociception.

The forebrain medial septum, which is an integral part of the septo-hippocampal network, is implicated in sensorimotor integration, fear and anxiety, and spatial learning and memory. A body of evidence also suggests that the septal region affects experimental pain. Indeed, some explorations in humans have raised the possibility that the region may modulate clinical pain as well. This review explores the evidence that implicates the medial septum in nociception and suggests that non-overlapping circuits in the region facilitate acute nociceptive behaviors and defensive behaviors that reflect affect and cognitive appraisal, especially in relation to persistent nociception. In line with a role in nociception, the region modulates nociceptive responses in the neuraxis, including the hippocampus and the anterior cingulate cortex. The aforementioned forebrain regions have also been implicated in persistent/long-lasting nociception. The review also weighs the effects of the medial septum on nociception vis-à-vis the known roles of the region and emphasizes the fact that the region is a part of network of forebrain structures which have been long associated with reward, cognition and affect-motivation and are now implicated in persistent/long-lasting nociception.

GABAergic neurons of the medial septum play a nodal role in facilitation of nociception-induced affect.

The present study explored the functional details of the influence of medial septal region (MSDB) on spectrum of nociceptive behaviours by manipulating intraseptal GABAergic mechanisms. Results showed that formalin-induced acute nociception was not affected by intraseptal microinjection of bicuculline, a GABAA receptor antagonist, or on selective lesion of septal GABAergic neurons. Indeed, the acute nociceptive responses were dissociated from the regulation of sensorimotor behaviour and generation of theta-rhythm by the GABAergic mechanisms in MSDB. The GABAergic lesion attenuated formalin-induced unconditioned cellular response in the anterior cingulate cortex (ACC) and blocked formalin-induced conditioned place avoidance (F-CPA), and as well as the contextual fear induced on conditioning with brief footshock. The effects of lesion on nociceptive-conditioned cellular responses were, however, variable. Interestingly, the lesion attenuated the conditioned representation of experimental context in dorsal hippocampus field CA1 in the F-CPA task. Collectively, the preceding suggests that the MSDB is a nodal centre wherein the GABAergic neurons mediate nociceptive affect-motivation by regulating cellular mechanisms in ACC that confer an aversive value to the noxious stimulus. Further, in conjunction with a modulatory influence on hippocampal contextual processing, MSDB may integrate affect with context as part of associative learning in the F-CPA task.