Increased interaction between endoplasmic reticulum and mitochondria following sleep deprivation.

BACKGROUND: Prolonged cellular activity may overload cell function, leading to high rates of protein synthesis and accumulation of misfolded or unassembled proteins, which cause endoplasmic reticulum (ER) stress and activate the unfolded protein response (UPR) to re-establish normal protein homeostasis. Previous molecular work has demonstrated that sleep deprivation (SD) leads to ER stress in neurons, with a number of ER-specific proteins being upregulated to maintain optimal cellular proteostasis. It is still not clear which cellular processes activated by sleep deprivation lead to ER- stress, but increased cellular metabolism, higher request for protein synthesis, and over production of oxygen radicals have been proposed as potential contributing factors. Here, we investigate the transcriptional and ultrastructural ER and mitochondrial modifications induced by sleep loss. RESULTS: We used gene expression analysis in mouse forebrains to show that SD was associated with significant transcriptional modifications of genes involved in ER stress but also in ER-mitochondria interaction, calcium homeostasis, and mitochondrial respiratory activity. Using electron microscopy, we also showed that SD was associated with a general increase in the density of ER cisternae in pyramidal neurons of the motor cortex. Moreover, ER cisternae established new contact sites with mitochondria, the so-called mitochondria associated membranes (MAMs), important hubs for molecule shuttling, such as calcium and lipids, and for the modulation of ATP production and redox state. Finally, we demonstrated that Drosophila male mutant flies (elav > linker), in which the number of MAMs had been genetically increased, showed a reduction in the amount and consolidation of sleep without alterations in the homeostatic sleep response to SD. CONCLUSIONS: We provide evidence that sleep loss induces ER stress characterized by increased crosstalk between ER and mitochondria. MAMs formation associated with SD could represent a key phenomenon for the modulation of multiple cellular processes that ensure appropriate responses to increased cell metabolism. In addition, MAMs establishment may play a role in the regulation of sleep under baseline conditions.

Characterization of Subcellular Organelles in Cortical Perisynaptic Astrocytes.

Perisynaptic astrocytic processes (PAPs) carry out several different functions, from metabolite clearing to control of neuronal excitability and synaptic plasticity. All these functions are likely orchestrated by complex cellular machinery that resides within the PAPs and relies on a fine interplay between multiple subcellular components. However, traditional transmission electron microscopy (EM) studies have found that PAPs are remarkably poor of intracellular organelles, failing to explain how such a variety of PAP functions are achieved in the absence of a proportional complex network of intracellular structures. Here, we use serial block-face scanning EM to reconstruct and describe in three dimensions PAPs and their intracellular organelles in two different mouse cortical regions. We described five distinct organelles, which included empty and full endosomes, phagosomes, mitochondria, and endoplasmic reticulum (ER) cisternae, distributed within three PAPs categories (branches, branchlets, and leaflets). The majority of PAPs belonged to the leaflets category (~60%), with branchlets representing a minority (~37%). Branches were rarely in contact with synapses (<3%). Branches had a higher density of mitochondria and ER cisternae than branchlets and leaflets. Also, branches and branchlets displayed organelles more frequently than leaflets. Endosomes and phagosomes, which accounted for more than 60% of all the organelles detected, were often associated with the same PAP. Likewise, mitochondria and ER cisternae, representing ~40% of all organelles were usually associated. No differences were noted between the organelle distribution of the somatosensory and the anterior cingulate cortex. Finally, the organelle distribution in PAPs did not largely depend on the presence of a spine apparatus or a pre-synaptic mitochondrion in the synapse that PAPs were enwrapping, with some exceptions regarding the presence of phagosomes and ER cisternae, which were slightly more represented around synapses lacking a spine apparatus and a presynaptic mitochondrion, respectively. Thus, PAPs contain several subcellular organelles that could underlie the diverse astrocytic functions carried out at central synapses.

Efficacy of a Combination of N-Palmitoylethanolamide, Beta-Caryophyllene, Carnosic Acid, and Myrrh Extract on Chronic Neuropathic Pain: A Preclinical Study.

Neuropathic pain (NP) is a common public health problem that poses a major challenge to basic scientists and health-care providers. NP is a complex problem with an unclear etiology and an often-inadequate response to current medications. Despite the high number of drugs available, their limited pharmacological efficacy and side effects hamper their chronic use. Thus, the search for novel treatments is a priority. In addition to pharmaceuticals, natural extracts and food supplements are often used to help treating patients with NP. One such supplement is Noxiall®, a commercially available combination of N-Palmitoylethanolamide (PEA), beta-caryophyllene; carnosic acid and myrrh. Here, we compare the efficacy of Noxiall® to that of the medications gabapentin and pregabalin in the NP model of chronic constriction injury (CCI) using sciatic nerve ligation in mouse. Following CCI, mice developed a significant increase in mechanical allodynia and thermal hyperalgesia. Results showed that administration of either Noxiall®, pregabalin, or gabapentin significantly attenuated mechanical allodynia. The magnitude of the Noxiall® effect was comparable to that of gabapentin or pregabalin. In addition, co-administration of non-effective doses of pregabalin and Noxiall® resulted in a significant decrease in NP, suggesting an additive efficacy. Noxiall® was efficacious also in reducing CCI-induced thermal hyperalgesia. These findings support the rationale of using natural remedies in conjunction with classical pharmacological agents to treat chronic NP.