N,N –Disubstituted piperazines and homopiperazines: Synthesis and affinities at α4β2* and α7* neuronal nicotinic acetylcholine receptors

A series of N, N– disubstituted piperazines and homopiperazines were prepared and evaluated for binding to natural α4β2* and α7* neuronal nicotinic acetylcholine receptors (nAChRs) using whole brain membrane. Some compounds exhibited good selectivity for α4β2* nAChRs and did not interact with the α7* nAChRs subtype. The most potent analogs were compounds 8-19 (K_i = 10.4 μM), 8–13 (K_i = 12.0 μM), and 8–24 (K_i = 12.8 μM). Thus, linking together a pyridine π-system and a cyclic amine moiety via a homopiperazine ring affords compounds with low affinity but with good selectivity for α4β2* nAChRs.     

Inactivation of Nitric Oxide by Uric Acid

The 1980 identification of nitric oxide (NO) as an endothelial cell-derived relaxing factor resulted in an unprecedented biomedical research of NO and established NO as one of the most important cardiovascular, nervous and immune system regulatory molecule. A reduction in endothelial cell NO levels leading to “endothelial dysfunction” has been identified as a key pathogenic event preceding the development of hypertension, metabolic syndrome, and cardiovascular disease. The reduction in endothelial NO in cardiovascular disease has been attributed to the action of oxidants that either directly react with NO or uncouple its substrate enzyme. In this report, we demonstrate that uric acid (UA), the most abundant antioxidant in plasma, reacts directly with NO in a rapid irreversible reaction resulting in the formation of 6-aminouracil and depletion of NO. We further show that this reaction occurs preferentially with NO even in the presence of oxidants peroxynitrite and hydrogen peroxide and that the reaction is at least partially blocked by glutathione. This study shows a potential mechanism by which UA may deplete NO and cause endothelial dysfunction, particularly under conditions of oxidative stress in which UA is elevated and intracellular glutathione is depleted.     

Cigarette smoke metabolically promotes cancer,via autophagy and premature aging in the host stromal microenvironment

Cigarette smoke has been directly implicated in the disease pathogenesis of a plethora of different human cancer subtypes, including breast cancers. The prevailing view is that cigarette smoke acts as a mutagen and DNA damaging agent in normal epithelial cells, driving tumor initiation. However, its potential negative metabolic effects on the normal stromal microenvironment have been largely ignored. Here, we propose a new mechanism by which carcinogen-rich cigarette smoke may promote cancer growth, by metabolically “fertilizing” the host microenvironment. More specifically, we show that cigarette smoke exposure is indeed sufficient to drive the onset of the cancer-associated fibroblast phenotype via the induction of DNA damage, autophagy and mitophagy in the tumor stroma. In turn, cigarette smoke exposure induces premature aging and mitochondrial dysfunction in stromal fibroblasts, leading to the secretion of high-energy mitochondrial fuels, such as L-lactate and ketone bodies. Hence, cigarette smoke induces catabolism in the local microenvironment, directly fueling oxidative mitochondrial metabolism (OXPHOS) in neighboring epithelial cancer cells, actively promoting anabolic tumor growth. Remarkably, these autophagic-senescent fibroblasts increased breast cancer tumor growth in vivo by up to 4-fold. Importantly, we show that cigarette smoke-induced metabolic reprogramming of the fibroblastic stroma occurs independently of tumor neo-angiogenesis. We discuss the possible implications of our current findings for the prevention of aging-associated human diseases and, especially, common epithelial cancers, as we show that cigarette smoke can systemically accelerate aging in the host microenvironment. Finally, our current findings are consistent with the idea that cigarette smoke induces the “reverse Warburg effect,” thereby fueling “two-compartment tumor metabolism” and oxidative mitochondrial metabolism in epithelial cancer cells.     

Prolactin receptor in regulation of neuronal excitability and channels

Prolactin (PRL) activates PRL receptor isoforms to exert regulation of specific neuronal circuitries, and to control numerous physiological and clinically-relevant functions including; maternal behavior, energy balance and food intake, stress and trauma responses, anxiety, neurogenesis, migraine and pain. PRL controls these critical functions by regulating receptor potential thresholds, neuronal excitability and/or neurotransmission efficiency. PRL also influences neuronal functions via activation of certain neurons, resulting in Ca²⁺ influx and/or electrical firing with subsequent release of neurotransmitters. Although PRL was identified almost a century ago, very little specific information is known about how PRL regulates neuronal functions. Nevertheless, important initial steps have recently been made including the identification of PRL-induced transient signaling pathways in neurons and the modulation of neuronal transient receptor potential (TRP) and Ca²⁺-dependent K⁺ channels by PRL. In this review, we summarize current knowledge and recent progress in understanding the regulation of neuronal excitability and channels by PRL.     

Glutathione transferase mu 2 protects glioblastoma cells against aminochrome toxicity by preventing autophagy and lysosome dysfunction

U373MG cells constitutively express glutathione S-transferase mu 2 (GSTM2) and exhibit ³H-dopamine uptake, which is inhibited by 2 µM of nomifensine and 15 µM of estradiol. We generated a stable cell line (U373MGsiGST6) expressing an siRNA against GSTM2 that resulted in low GSTM2 expression (26% of wild-type U373MG cells). A significant increase in cell death was observed when U373MGsiGST6 cells were incubated with 50 µM purified aminochrome (18-fold increase) compared with wild-type cells. The incubation of U373MGsiGST6 cells with 75 µM aminochrome resulted in the formation of autophagic vacuoles containing undigested cellular components, as determined using transmission electron microscopy. A significant increase in autophagosomes was determined by measuring endogenous LC3-II, a significant decrease in cell death was observed in the presence of bafilomycin A₁, and a significant increase in cell death was observed in the presence of trehalose. A significant increase in LAMP2 immunostaining was observed, a significant decrease in bright red fluorescence of lysosomes with acridine orange was observed, and bafilomycin A₁ pretreatment reduced the loss of lysosome acidity. A significant increase in cell death was observed in the presence of lysosomal protease inhibitors. Aggregation of TUBA/α-tubulin (tubulin, α) and SQSTM1 protein accumulation were also observed. Moreover, a significant increase in the number of lipids droplets was observed compared with U373MG cells with normal expression of GSTM2. These results support the notion that GSTM2 is a protective enzyme against aminochrome toxicity in astrocytes and that aminochrome cell death in U373MGsiGST6 cells involves autophagic-lysosomal dysfunction.     

Acid Phosphatase as a Selective Marker for a Class of Small Sensory Ganglion Cells in Several Mammals: Spinal Cord Distribution,Histochemical Properties,and Relation to Fluoride-Resistant Acid Phosphatase (FRAP) of Rodents

Fluoride-resistant acid phosphatase (FRAP) activity as characterized in rat and mouse was studied in sensory ganglion and spinal cord of several mammals, using both the Gomori lead-ion capture and azo-dye coupling methods. FRAP was specifically localized to small- and medium-diameter primary afferent neurons and inner substantia gelatinosa of all nonrodent animals studied, including rabbit, cat, dog, monkey, cow, and human. In rabbit, sciatic nerve transection resulted in depletion of enzymatic activity in ipsilateral spinal cord dorsal horn in a pattern corresponding to the distribution of central terminals of the nerve. Further analysis of the substrate specificity and pH dependence of FRAP was carried out primarily in rat sensory ganglion and spinal cord; the enzyme was found to hydrolyze a wide variety of phosphomonoesters in a relatively nonselective manner at both pH 5 and pH 7, including 5′-nucleotides, phosphorylated amino acids, and several exogenous compounds.

The visualization of FRAP-like activity in several nonrodent species is discussed with reference to previous work indicating its presence only in mouse and rat. Technical factors are considered that limit the applicability of the lead-ion histochemical method in demonstration of FRAP and in efforts at functional characterization of the enzyme, especially in light of its ability to hydrolyze a broad spectrum of substrates over a wide pH range. Alternative interpretations of the expression of acid phosphatase activity in a select class of small sensory ganglion cells are suggested, including several possible nonsynaptic roles of FRAP in the peripheral nervous system.     

Joint distribution of first exit times of a two dimensional Wiener process with jumps with application to a pair of coupled neurons

Motivated by a neuronal modeling problem, a bivariate Wiener process with two independent components is considered. Each component evolves independently until one of them reaches a threshold value. If the first component crosses the threshold value, it is reset while the dynamics of the other component remains unchanged. But, if this happens to the second component, the first one has a jump of constant amplitude; the second component is then reset to its starting value and its evolution restarts. Both processes evolve once again until one of them reaches again its boundary. In this work, the coupling of the first exit times of the two connected processes is studied.     

Growth Arrest Specific 1 (GAS1) Is Abundantly Expressed in the Adult Mouse Central Nervous System

Growth arrest specific 1 (GAS1) is a pleiotropic protein that induces apoptosis and cell arrest in different tumors, but it is also involved in the development of the nervous system and other tissues and organs. This dual ability is likely caused by its capacity to interact both by inhibiting the intracellular signaling cascade induced by glial cell-line derived neurotrophic factor and by facilitating the activity of the sonic hedgehog pathway. The presence of GAS1 mRNA has been described in adult mouse brain, and here we corroborated this observation. We then proceeded to determine the distribution of the protein in the adult central nervous system (CNS). We detected, by western blot analysis, expression of GAS1 in olfactory bulb, caudate-putamen, cerebral cortex, hippocampus, mesencephalon, medulla oblongata, cerebellum, and cervical spinal cord. To more carefully map the expression of GAS1, we performed double-label immunohistochemistry and noticed expression of GAS1 in neurons in all brain areas examined. We also observed expression of GAS1 in astroglial cells, albeit the pattern of expression was more restricted than that seen in neurons. Briefly, in the present article, we report the widespread distribution and cellular localization of the GAS1 native protein in adult mammalian CNS.