Nitric oxide synthase inhibitor attenuates the effects of repeated restraint stress on synaptic transmission in the paraventricular nucleus of the rat hypothalamus

Corticotropin-releasing hormone (CRH)-synthesizing parvocellular neuroendocrine cells (PNCs) of the hypothalamic paraventricular nucleus (PVN) play a key role in the activation of the hypothalamic-pituitary-adrenocortical (HPA) axis. Several studies have demonstrated that synaptic inputs to these cells may undergo stress-related enhancement but, on the other hand, it has been reported that exposition to the same stressor for prolonged time periods may induce a progressive reduction in the response of the HPA axis to homotypic stressors. In the present study rats were subjected to 10 min restraint sessions, repeated twice daily for 3 or 7 days. Miniature excitatory and inhibitory postsynaptic currents (mEPSCs and mIPSCs) were then recorded from PNCs in ex vivo hypothalamic slice preparations obtained 24 h after the last restraint. Restraint stress repeated over 3 days resulted in increased mean frequency and decreased rise time and decay time constant of mEPSCs, accompanied by a decrease in the excitability of PNCs, however, no such changes were evident in slices obtained from rats subjected to restraint over 7 days. There were no changes in mIPSCs after repeated restraint. Administration of the unspecific nitric oxide synthase (NOS) blocker Nω-Nitro-L-arginine (L-NNA) before each restraint, repeated over 3 days, prevented the occurrence of an increase in mEPSC frequency. However, animals receiving L-NNA and subjected to repeated restraint had similar changes in PNCs membrane excitability and mEPSC kinetics as stressed rats not receiving L-NNA. Comparison of the effects of a single 10 min restraint session followed by either an immediate or delayed (24 h) decapitation revealed an increase in the mean mEPSC frequency and a decrease in the mean mIPSC frequency in slices prepared immediately after restraint, with no apparent effects when slice preparation was delayed by 24 h. These results demonstrate that restraint, lasting 10 min and repeated twice daily for 3 days, induces a selective and long-lasting enhancement of excitatory synaptic input onto PNCs, partially by a NOS-dependent mechanism, and reduces PNC excitability, whereas prolongation of repeated stress for up to 7 days results in an adaptation

Mitochondria-associated membranes in aging and senescence: Structure, function, and dynamics

Sites of close contact between mitochondria and the endoplasmic reticulum (ER) are known as mitochondria-associated membranes (MAM) or mitochondria-ER contacts (MERCs), and play an important role in both cell physiology and pathology. A growing body of evidence indicates that changes observed in the molecular composition of MAM and in the number of MERCs predisposes MAM to be considered a dynamic structure. Its involvement in processes such as lipid biosynthesis and trafficking, calcium homeostasis, reactive oxygen species production, and autophagy has been experimentally confirmed. Recently, MAM have also been studied in the context of different pathologies, including Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, type 2 diabetes mellitus and GM1-gangliosidosis. An underappreciated amount of data links MAM with aging or senescence processes. In the present review, we summarize the current knowledge of basic MAM biology, composition and action, and discuss the potential connections supporting the idea that MAM are significant players in longevity

Interaction of mitochondria with the endoplasmic reticulum and plasma membrane in calcium homeostasis, lipid trafficking and mitochondrial structure

Studying organelles in isolation has been proven to be indispensable for deciphering the underlying mechanisms of molecular cell biology. However, observing organelles in intact cells with the use of microscopic techniques reveals a new set of different junctions and contact sites between them that contribute to the control and regulation of various cellular processes, such as calcium and lipid exchange or structural reorganization of the mitochondrial network. In recent years, many studies focused their attention on the structure and function of contacts between mitochondria and other organelles. From these studies, findings emerged showing that these contacts are involved in various processes, such as lipid synthesis and trafficking, modulation of mitochondrial morphology, endoplasmic reticulum (ER) stress, apoptosis, autophagy, inflammation and Ca2+handling. In this review, we focused on the physical interactions of mitochondria with the endoplasmic reticulum and plasma membrane and summarized present knowledge regarding the role of mitochondria-associated membranes in calcium homeostasis and lipid metabolism

Apparent Anomalous Diffusion in the Cytoplasm of Human Cells: The Effect of Probes’ Polydispersity

This work, based on <i>in vivo</i> and <i>in vitro</i> measurements, as well as <i>in silico</i> simulations, provides a consistent analysis of diffusion of polydisperse nanoparticles in the cytoplasm of living cells. Using the example of fluorescence correlation spectroscopy (FCS), we show the effect of polydispersity of probes on the experimental results. Although individual probes undergo normal diffusion, in the ensemble of probes, an effective broadening of the distribution of diffusion times occurssimilar to anomalous diffusion. We introduced fluorescently labeled dextrans into the cytoplasm of HeLa cells and found that cytoplasmic hydrodynamic drag, exponentially dependent on probe size, extraordinarily broadens the distribution of diffusion times across the focal volume. As a result, the <i>in vivo</i> FCS data were effectively fitted with the anomalous subdiffusion model while for a monodisperse probe the normal diffusion model was most suitable. Diffusion time obtained from the anomalous diffusion model corresponds to a probe whose size is determined by the weight-average molecular weight of the polymer. The apparent anomaly exponent decreases with increasing polydispersity of the probes. Our results and methodology can be applied in intracellular studies of the mobility of nanoparticles, polymers, or oligomerizing proteins