Sildenafil prevents right ventricular hypertrophy and improves heart rate variability in rats with pulmonary hypertension secondary to experimental diabetes.

Chronic treatment with sildenafil (SILD) is an effective protector on the development of cardiovascular complications of pulmonary hypertension (PH) and diabetes. However, to date, no studies have evaluated the effect of SILD on cardiopulmonary pathophysiology during PH secondary to type 1 diabetes. AIM: The present study aimed to evaluate the beneficial effects of chronic SILD treatment on pulmonary arterial pressure, right ventricular hypertrophy (RVH) and cardiac autonomic dysfunction in rats with PH secondary to diabetes. METODOLOGY: Male Sprague Dawley rats were randomly distributed into the control group (saline), diabetic group (60 mg/kg with streptozotocin), SILD-treated control group (20 mg/kg) and SILD-treated diabetic group. RESULTS: After 8 weeks the type 1 diabetic animals presented PH, endothelial dysfunction of the pulmonary arteries, electrocardiographic alterations, RVH and overexpression of phosphodiesterase type 5 in the heart. In type 1 diabetic animals, SILD treatment prevented the development of PH, endothelial dysfunction and RVH. SILD treatment also prevented alterations in the corrected QT period and heart rate variability and prevented overexpression of phosphodiesterase type 5. CONCLUSION: Our results indicate for the first time that SILD treatment prevents pulmonary arterial endothelial dysfunction, pulmonary hypertension, right ventricular hypertrophy and improves heart rate variability in type 1 diabetic rats.

Convergent evidence from alcohol-dependent humans and rats for a hyperdopaminergic state in protracted abstinence.

A major hypothesis in addiction research is that alcohol induces neuroadaptations in the mesolimbic dopamine (DA) system and that these neuroadaptations represent a key neurochemical event in compulsive drug use and relapse. Whether these neuroadaptations lead to a hypo- or hyperdopaminergic state during abstinence is a long-standing, unresolved debate among addiction researchers. The answer is of critical importance for understanding the neurobiological mechanism of addictive behavior. Here we set out to study systematically the neuroadaptive changes in the DA system during the addiction cycle in alcohol-dependent patients and rats. In postmortem brain samples from human alcoholics we found a strong down-regulation of the D1 receptor- and DA transporter (DAT)-binding sites, but D2-like receptor binding was unaffected. To gain insight into the time course of these neuroadaptations, we compared the human data with that from alcohol-dependent rats at several time points during abstinence. We found a dynamic regulation of D1 and DAT during 3 wk of abstinence. After the third week the rat data mirrored our human data. This time point was characterized by elevated extracellular DA levels, lack of synaptic response to D1 stimulation, and augmented motor activity. Further functional evidence is given by a genetic rat model for hyperdopaminergia that resembles a phenocopy of alcohol-dependent rats during protracted abstinence. In summary, we provide a new dynamic model of abstinence-related changes in the striatal DA system; in this model a hyperdopaminergic state during protracted abstinence is associated with vulnerability for relapse.

Activation of 5-HT receptors inhibits GABAergic transmission by pre-and post-synaptic mechanisms in layer II/III of the juvenile rat auditory cortex.

The specific mechanisms by which serotonin (5-HT) modulates synaptic transmission in the auditory cortex are still unknown. In this work, we used whole-cell recordings from layer II/III of pyramidal neurons in rat brain slices to characterize the influence of 5-HT on inhibitory synaptic activity in the auditory cortex after pharmacological blockade of excitatory glutamatergic transmission. We found that bath application of 5-HT (5 µM) reduced the frequency and amplitude of both spontaneous and miniature inhibitory postsynaptic currents (IPSCs), reduced the amplitude of evoked IPSCs, and enhanced facilitation of paired pulse ratio (PPR), suggesting presynaptic inhibition. To determine which the serotonin receptors were involved in this effect, we studied the influence of specific 5-HT receptor agonists and antagonists on É£-aminobutyric acid (GABA)ergic synaptic transmission. The inhibiting influence of 5-HT in the GABAergic synaptic activity was mimicked by using the selective agonists of the 5-HT1A and 5-HT2A receptors, 8(OH)-DPAT (10 µM) and DOI (10 µM), respectively; and it was prevented by their respective antagonists NAN-190 (1 µM) and ritanserin (1 µM). Furthermore, the application of the selective agonist of 5-HT1A receptors, 8-(OH)-DPAT (10 µM), produced PPR facilitation, while DOI application (5-HT2A agonist) did not change the PPR. Moreover, the 5-HT2A agonist reduced the amplitude of the IPSCs evoked by application of the selective GABA agonist, muscimol. These results suggest a presynaptic and postsynaptic reduction of GABAergic transmission mediated by 5-HT1A and 5-HT2A serotonergic receptors, respectively.

Computational Molecular Characterization of the Interaction of Acetylcholine and the NMDA Receptor to Explain the Direct Glycine-Competitive Potentiation of NMDA-Mediated Neuronal Currents.

The activation of N-methyl-d-aspartate receptor (NMDAR) is triggered by the closure of bilobed (D1 and D2) clamshell-like clefts upon binding glycine (Gly) and glutamate. There is evidence that cholinergic compounds modulate NMDAR-mediated currents via direct receptor-ligand interactions; however, molecular bases are unknown. Here, we first propose a mechanistic structure-based explanation for the observed ACh-induced submaximal potentiation of NMDA-elicited currents in striatal neurons by predicting competitive inhibition with Gly. Then, the model was validated, in principle, by confirming that the coapplication of Gly and ACh significantly reduces these neuronal currents. Finally, we delineate the interplay of ACh with the NMDAR by a combination of computational strategies. Crystallographic ACh-bound complexes were studied, revealing a similar ACh binding environment on the GluN1 subunit of the NMDAR. We illustrate how ACh can occupy X-ray monomeric open, dimeric "semiopen" cleft conformations obtained by molecular dynamics and a full-active cryo-EM NMDAR structure, explaining the suboptimal NMDAR electrophysiological activity under the "Venus Flytrap model". At an evolutionary biology level, the binding mode of ACh coincides with that of the homologous ornithine-bound periplasmic LAO binding protein complex. Our computed results indicate an analogous mechanism of action, inasmuch as ACh may stabilize the GluN1 subunit "semiclosed" conformations by inducing direct and indirect D1-to-D2 interdomain bonds. Additionally, an alternative binding site was detected, shared by the known NMDAR allosteric modulators. Experimental and computed results strongly suggest that ACh acts as a Gly-competitive, submaximal potentiating agent of the NMDAR, possibly constituting a novel chemotype for multitarget-directed drug development, e.g., to treat Alzheimer's, and it may lead to a new understanding of glutamatergic neurotransmission.

Increased Reliance on Carbohydrates for Aerobic Exercise in Highland Andean Leaf-Eared Mice, but Not in Highland Lima Leaf-Eared Mice.

Exercise is an important performance trait in mammals and variation in aerobic capacity and/or substrate allocation during submaximal exercise may be important for survival at high altitude. Comparisons between lowland and highland populations is a fruitful approach to understanding the mechanisms for altitude differences in exercise performance. However, it has only been applied in very few highland species. The leaf-eared mice (LEM, genus Phyllotis) of South America are a promising taxon to uncover the pervasiveness of hypoxia tolerance mechanisms. Here we use lowland and highland populations of Andean and Lima LEM (P. andium and P. limatus), acclimated to common laboratory conditions, to determine exercise-induced maximal oxygen consumption (V˙O2max), and submaximal exercise metabolism. Lowland and highland populations of both species showed no difference in V˙O2max running in either normoxia or hypoxia. When run at 75% of V˙O2max, highland Andean LEM had a greater reliance on carbohydrate oxidation to power exercise. In contrast, highland Lima LEM showed no difference in exercise fuel use compared to their lowland counterparts. The higher carbohydrate oxidation seen in highland Andean LEM was not explained by maximal activities of glycolytic enzymes in the gastrocnemius muscle, which were equivalent to lowlanders. This result is consistent with data on highland deer mouse populations and suggests changes in metabolic regulation may explain altitude differences in exercise performance.

Population history, phylogeography, and conservation genetics of the last Neotropical mega-herbivore, the lowland tapir (Tapirus terrestris).

BACKGROUND: Understanding the forces that shaped Neotropical diversity is central issue to explain tropical biodiversity and inform conservation action; yet few studies have examined large, widespread species. Lowland tapir (Tapirus terrrestris, Perissodactyla, Tapiridae) is the largest Neotropical herbivore whose ancestors arrived in South America during the Great American Biotic Interchange. A Pleistocene diversification is inferred for the genus Tapirus from the fossil record, but only two species survived the Pleistocene megafauna extinction. Here, we investigate the history of lowland tapir as revealed by variation at the mitochondrial gene Cytochrome b, compare it to the fossil data, and explore mechanisms that could have shaped the observed structure of current populations. RESULTS: Separate methodological approaches found mutually exclusive divergence times for lowland tapir, either in the late or in the early Pleistocene, although a late Pleistocene divergence is more in tune with the fossil record. Bayesian analysis favored mountain tapir (T. pinchaque) paraphyly in relation to lowland tapir over reciprocal monophyly, corroborating the inferences from the fossil data these species are sister taxa. A coalescent-based analysis rejected a null hypothesis of allopatric divergence, suggesting a complex history. Based on the geographic distribution of haplotypes we propose (i) a central role for western Amazonia in tapir diversification, with a key role of the ecological gradient along the transition between Andean subcloud forests and Amazon lowland forest, and (ii) that the Amazon river acted as an barrier to gene flow. Finally, the branching patterns and estimates based on nucleotide diversity indicate a population expansion after the Last Glacial Maximum. CONCLUSIONS: This study is the first examining lowland tapir phylogeography. Climatic events at the end of the Pleistocene, parapatric speciation, divergence along the Andean foothill, and role of the Amazon river, have similarly shaped the history of other taxa. Nevertheless further work with additional samples and loci is needed to improve our initial assessment. From a conservation perspective, we did not find a correspondence between genetic structure in lowland tapir and ecogeographic regions proposed to define conservation priorities in the Neotropics. This discrepancy sheds doubt into this scheme's ability to generate effective conservation planning for vagile species.

Noisy Light Augments the Na+ Current in Somatosensory Pyramidal Neurons of Optogenetic Transgenic Mice.

In previous reports, we developed a method to apply Brownian optogenetic noise-photostimulation (BONP, 470 nm) up to 0.67 mW on the barrel cortex of in vivo ChR2 transgenic mice. In such studies, we found that the BONP produces an increase in the evoked field potentials and the neuronal responses of pyramidal neurons induced by somatosensory mechanical stimulation. Here we extended such findings by examining whether the same type of BONP augments the Na+ current amplitude elicited by voltage-clamp ramps of dissociated pyramidal neurons from the somatosensory cortex of ChR2 transgenic and wild type mice. We found that in all neurons from the ChR2 transgenic mice, but none of the wild type mice, the peak amplitude of a TTX-sensitive Na+ current and its inverse of latency exhibited inverted U-like graphs as a function of the BONP level. It means that an intermediate level of BONP increases both the peak amplitude of the Na+ current and its inverse of latency. Our research suggests that the impact of BONP on the Na+ channels of pyramidal neurons could be associated with the observed augmentation-effects in our previous in vivo preparation. Moreover, it provides caution information for the use of an appropriate range of light intensity, <0.67 mW, which could avoid opto non-genetics (also termed "optonongenetic") related responses due to light-induced temperature changes.

Cholinergic direct inhibition of N-methyl-D aspartate receptor-mediated currents in the rat neocortex.

Acetylcholine (ACh) and N-methyl-D aspartate receptors (NMDARs) interact in the regulation of multiple important brain functions. NMDAR activation is indirectly modulated by ACh through the activation of muscarinic or nicotinic receptors. Scant information is available on whether ACh directly interacts with the NMDAR. By using a cortical brain slice preparation we found that the application of ACh and of other drugs acting on muscarinic or nicotinic receptors induces an acute and reversible reduction of NMDAR-mediated currents (I(NMDA)), ranging from 20 to 90% of the control amplitude. The reduction displayed similar features in synaptic I(NMDA) in brain slices, as well as in currents evoked by NMDA application in brain slices or from acutely dissociated cortical cells, demonstrating its postsynaptic nature. The cholinergic inhibition of I(NMDA) displayed an onset-offset rate in the order of a second, and was resistant to the presence of the muscarinic antagonist atropine (10 microM) in the extracellular solution, and of G-protein blocker GDP(beta)S (500 microM) and activator GTP(gamma)S (400 microM) in the intracellular solution, indicating that it was not G-protein dependent. Recording at depolarized or hyperpolarized holding voltages reduced NMDAR-mediated currents to similar extents, suggesting that the inhibition was voltage-independent, whereas the reduction was markedly more pronounced in the presence of glycine (20 microM). A detailed analysis of the effects of tubocurarine suggested that at least this drug interfered with glycine-dependent NMDAR-activity. We conclude that NMDAR-mediated current scan be inhibited directly by cholinergic drugs, possibly by direct interaction within one or more subunits of the NMDAR. Our results could supply a new interpretation to previous studies on the role of ACh at the glutamatergic synapse.

Effects of Short-Term Random Noise Electrical Stimulation on Dissociated Pyramidal Neurons from the Cerebral Cortex.

Transcranial random noise electrical stimulation (tRNS) of the human brain is a non-invasive technique that can be employed to increase the excitability of the cerebral cortex; however, the physiological mechanisms remain unclear. Here we report for the first time the effects of short-term (250 ms) random noise electrical stimulation (RNS) on in-vitro acutely-isolated brain pyramidal neurons from the somatosensory and auditory cerebral cortex. We analyzed the correlation between the peak amplitude of the Na+ current and its latency for different levels of RNS. We found three groups of neurons. The first group exhibited a positive correlation, the second, a negative correlation, and the third group of neurons did not exhibit correlation. In the first group, both the peak amplitude of a TTX-sensitive Na+ current and its inverse of latency followed similar inverted U-like functions relative to the electrical RNS level. In this group, the RNS levels in which the maximal values of the inverted U-like functions occurred were the same. In the second group, the maximal values of the inverted U-like functions occurred at different levels. In the third group, only the peak amplitude of the Na+ current exhibited a clear inverted U-like function, but the inverse of the latency versus the electrical RNS, did not exhibit a clear inverted U-like function. A Hodgkin-Huxley neuron model reproduces our experimental results and shows that the observed behavior in the Na+ current could be due to the impact of RNS on the kinetics of activation and inactivation of the Na+ channels.

Increase in carbohydrate utilization in high-altitude Andean mice.

The low oxygen levels at high altitude are a potent and unavoidable physiological stressor to which highland mammals must adapt. One hypothesized adaptation to high altitude is an increased reliance on carbohydrates to support aerobic activities. Based on stoichiometries of combustion, ATP yield per mole of oxygen from carbohydrates is approximately 15% higher than from lipids (observed difference closer to 30%), and increased carbohydrate use represents an important oxygen-saving strategy that may be under high selective pressure. Although this hypothesis was first proposed nearly 30 years ago, the in vivo patterns of whole-body fuel use during exercise remain undefined for any highland mammal (including humans). Here we use a powerful multispecies approach to show that wild-caught high-altitude (4,000-4,500 m) native species of mice (Phyllotis andium and Phyllotis xanthopygus) from the Peruvian Andes use proportionately more carbohydrates and have higher oxidative capacities of cardiac muscles compared to closely related low-altitude (100-300 m) native counterparts (Phyllotis amicus and Phyllotis limatus). These results strongly infer that highland Phyllotis have evolved a metabolic strategy to economize oxygen when performing energy-demanding tasks at altitude. This study provides compelling evidence of adjustments in fuel use as an adaptation to high-altitude hypoxia in mammals.

Chromosomal aberrations in peripheral lymphocytes from male native miners working in the Peruvian Andes

We analyzed the frequency of chromosomal aberrations in peripheral lymphocytes from underground miners from the Casapalca (n = 8, mean age = 45 y, range = 36 y to 55 y) and Bellavista (n = 8, mean age = 28 y, range 23 y to 34 y) high-altitude mining camps in the Peruvian Andes. This population was occupationally exposed to heavy metals such as lead and zinc as well as diesel emission particles, organic solvents and mine dust. The control groups consisted of individuals from a high altitude farming community in the Peruvian village of Tinco (n = 8, mean age = 37 y, range = 25 y to 52 y) and the sea level city of Lima (n = 14, mean age = 26 y, range = 20 y to 35 y). All individuals were male native Peruvians. A significantly higher incidence (1.88 percent, p < 0.05) of chromosomal aberrations (chromatid deletions and breaks, chromosome breaks and acentric fragments) were detected in lymphocytes from miners at the Casapalca camp as compared to miners from the Bellavista camp (0.5 percent, chromatid deletions and acentric fragments only) and the Lima sea level (0.07 percent, chromatid deletions only) and Tinco high altitude (no aberrations) controls. These results suggest that male native Peruvians occupationally exposed to underground mining activity have an increased frequency of chromosomal aberrations, which could be related to both age and exposure time. The increased chromosomal damage observed in the mining populations studied may be attributable to the complex mixture of genotoxic agents to which the miners may have been exposed.