Frederico C. Pereira

Early cardiac changes in a rat model of prediabetes: brain natriuretic peptide overexpression seems to be the best marker

BackgroundDiabetic cardiomyopathy (DCM) is defined as structural and functional changes in the myocardium due to metabolic and cellular abnormalities induced by diabetes mellitus (DM). The impact of prediabetic conditions on the cardiac tissue remains to be elucidated. The goal of this study was to elucidate whether cardiac dysfunction is already present in a state of prediabetes, in the presence of insulin resistance, and to unravel the underlying mechanisms, in a rat model without obesity and hypertension as confounding factors.MethodsTwo groups of 16-week-old Wistar rats were tested during a 9 week protocol: high sucrose (HSu) diet group (n = 7) – rats receiving 35% of sucrose in drinking water vs the vehicle control group (n = 7). The animal model was characterized in terms of body weight (BW) and the glycemic, insulinemic and lipidic profiles. The following parameters were assessed to evaluate possible early cardiac alterations and underlying mechanisms: blood pressure, heart rate, heart and left ventricle (LV) trophism indexes, as well as the serum and tissue protein and/or the mRNA expression of markers for fibrosis, hypertrophy, proliferation, apoptosis, angiogenesis, endothelial function, inflammation and oxidative stress.ResultsThe HSu-treated rats presented normal fasting plasma glucose (FPG) but impaired glucose tolerance (IGT), accompanied by hyperinsulinemia and insulin resistance (P < 0.01), confirming this rat model as prediabetic. Furthermore, although hypertriglyceridemia (P < 0.05) was observed, obesity and hypertension were absent. Regarding the impact of the HSu diet on the cardiac tissue, our results indicated that 9 weeks of treatment might be associated with initial cardiac changes, as suggested by the increased LV weight/BW ratio (P < 0.01) and a remarkable brain natriuretic peptide (BNP) mRNA overexpression (P < 0.01), together with a marked trend for an upregulation of other important mediators of fibrosis, hypertrophy, angiogenesis and endothelial lesions, as well as oxidative stress. The inflammatory and apoptotic markers measured were unchanged.ConclusionsThis animal model of prediabetes/insulin resistance could be an important tool to evaluate the early cardiac impact of dysmetabolism (hyperinsulinemia and impaired glucose tolerance with fasting normoglycemia), without confounding factors such as obesity and hypertension. Left ventricle hypertrophy is already present and brain natriuretic peptide seems to be the best early marker for this condition.

May Exercise Prevent Addiction

Amphetamines exert their persistent addictive effects by activating brains reward pathways, perhaps through the release of dopamine in the nucleus accumbens (and/or in other places). On the other hand, there is a relationship between dopamine and all behavioural aspects that involve motor activity and it has been demonstrated that exercise leads to an increase in the synthesis and release of dopamine, stimulates neuroplasticity and promotes feelings of well-being. Moreover, exercise and drugs of abuse activate overlapping neural systems. Thus, our aim was to study the influence of chronic exercise in the mechanism of addiction using an amphetamine-induced conditioned-place-preference in rats. Adult male Sprague-Dawley rats were randomly separated in groups with and without chronic exercise. Chronic exercise consisted in a 8 week treadmill running program, with increasing intensity. The conditioned place preference test was performed in both groups using a procedure and apparatus previously established. A 2 mg.kg-1 amphetamine or saline solution was administered intraperitonially according to the schedule of the conditioned place preference. Before conditioning none of the animals showed preference for a specific compartment of the apparatus. The used amphetamine dose in the conditioning phase was able to produce a marked preference towards the drug-associated compartment in the group without exercise. In the animals with exercise a significant preference by the compartment associated with saline was observed. These results lead us to conclude that a previous practice of regular physical activity may help preventing amphetamine addiction in the conditions used in this test.

Methamphetamine Changes NMDA and AMPA Glutamate Receptor Subunit Levels in the Rat Striatum and Frontal Cortex

Methamphetamine (METH) is a powerful psychostimulant whose noxious effects depend largely on the pattern of abuse. METH‐induced glutamate release may overactivate N‐methyl‐d‐aspartate and α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionic acid receptors (NMDAR and AMPAR, respectively) causing excitotoxicity. In the brain, these receptors are also known for their essential role in mediating memory consolidation. Therefore, we assessed glial fibrillary acidic protein (GFAP) expression as a marker for astrogliosis and neurodegeneration by using Fluoro‐Jade C (F‐J C) staining. Moreover, we investigated the effect of two METH regimens on NMDAR NR1 and NR2A and on AMPAR GluR2 subunit expression in the rat striatum and frontal cortex 24 h after drug treatment. Adult Sprague‐Dawley rats were injected subcutaneously (s.c.) on six consecutive days with saline (control and acute groups) or with an increasing dose of METH (10, 15, 15, 20, 20, 25 mg/kg/day; ED group). On the seventh day, both METH groups were given a “bolus” of 30 mg/kg METH, whereas controls received saline. We evaluated the expression levels of GFAP by both Western blot and immunohistochemical assays and concluded that there was no difference from control levels. In addition, neither drug regimen resulted in neurodegeneration within 24 h of last METH administration. In the frontal cortex of the acute group, NR1 expression level was decreased, and both NR2A and GluR2 were increased. Also, in the striatum of the acute group, the expression level of GluR2 was significantly increased, and both GluR2 and NR2A levels were augmented in the striatum of the ED group. Taken together, these results suggest a protective mechanism by decreasing permeability and/or functionality of AMPAR and NMDAR to counteract METH‐induced glutamate overflow in the brain. Moreover, these results may explain, in part, the mnemonic deficits and psychotic behavior associated with METH abuse.

Methamphetamine, morphine, and their combination: acute changes in striatal dopaminergic transmission evaluated by microdialysis in awake rats.

Abstract:  The co‐administration of methamphetamine (METH) and MOR (MOR)‐like compounds is becoming increasingly popular among drug abusers. Recently, it was demonstrated that rats would self‐inject METH–heroin combination and that this combination produced a greater rewarding effect than the identical doses of METH alone and it was further suggested that enhanced reward might underlie the popularity of this combination. However, there is null information on the effects of the MOR–METH combination on striatal dopaminergic transmission. In the present article, in vivo brain microdialysis was used to examine the effects of two METH doses (1 and 5 mg/kg, i.p.; [METH1: hyperlocomotion‐inducing] and [METH5: stereotypy‐inducing], respectively) and MOR (10 mg/kg, i.p. [MOR10]) either alone or in combination on dopamine (DA) and 3,4‐dihydroxyphenylacetic acid (DOPAC) release in caudate putamen (CPu) in freely moving rats. METH1 evoked a transient threefold increase in DA overflow in only one‐third of dosed rats. On the contrary, METH5 elicited a 11‐fold increase in the extracellular DA levels 30 min after dosing and stayed significantly (P < 0.05) above control levels up to 1.5 h. On the other hand, MOR10 did not significantly change DA extracellular levels. MOR10–METH1 combination prolonged DA outflow for 1 h in all rats dosed without changing peak effect compared to METH1. On the other hand, MOR10–METH5 combination did not change the peak effect nor the DA outflow profile compared to METH5 alone. Consistently, there is a concentration‐dependent decrease in DOPAC efflux evoked by METH: METH1 evoked a smaller decrease in DOPAC outflow showing a tendency for returning to basal values whereas METH5 kept DOPAC extracellular levels reduced throughout the experiment. Again, MOR10 did not significantly change DOPAC extracellular levels. MOR delayed the onset without changing METH effect on the DOPAC output. These findings provide suggestive evidence that MOR potentiated the increase in extracellular DA levels induced by a low dose of METH. Thus, this combination yields a profile of action that might underlie the reinforcing properties sought by drug addicts.

Influence of Chronic Exercise on the Amphetamine-Induced Dopamine Release and Neurodegeneration in the Striatum of the Rat

The aim of this study was to verify the effect of chronic exercise on the striatal dopamine (DA) outflow induced by low and high single doses of amphetamine (AMPH), and verify the existence of an exercise protective role on neurodegeneration. Adult male Sprague‐Dawley rats were randomly separated into six groups: chronic exercise, saline; chronic exercise, 5 mg kg−1 AMPH; chronic exercise, 30 mg kg−1 AMPH; without exercise, saline; without exercise, 5 mg kg−1 AMPH; without exercise, 30 mg kg−1 AMPH. Chronic exercise consisted of an 8‐week running program on a treadmill, with increasing intensity. Animals were anesthetized, placed into a stereotaxic frame and an intracerebral guide cannula implanted into the caudate–putamen. When indicated, microdialysis was performed. Dialysate samples were collected during 30‐min intervals for 6 h, before and after the intraperitonial administration of AMPH or saline solution. HPLC with electrochemical detection was used to quantify DA. Chronic exercise did not significantly change the extracellular DA basal values. Regarding the maximal DA levels in the dialysates, in the rats treated with 5 mg kg−1 AMPH, there was no significant difference between groups with and without chronic exercise; on the contrary, in animals treated with 30 mg kg−1 AMPH, the DA release was lower in the group with chronic exercise. Moreover, the maintenance of higher levels of DA along time in the training group suggests a diminished reuptake of DA. By using the Fluoro‐Jade C staining technique, we did not find neuronal death in any of the groups. In conclusion, these results suggest that chronic exercise leads to a diminished release and reuptake of DA after administration of a high dose of AMPH, whereas neither chronic exercise nor AMPH seems to induce neurodegeneration.

Disruption of striatal glutamatergic/GABAergic homeostasis following acute methamphetamine in mice

Methamphetamine leads to functional changes in basal ganglia that are linked to impairment in motor activity. Previous studies from our group and others have shown that a single high-methamphetamine injection induces striatal dopaminergic changes in rodents. However, striatal glutamatergic, GABAergic and serotoninergic changes remain elusive under this methamphetamine regimen. Moreover, nothing is known about the participation of the receptor for advanced glycation end-products (RAGE), which is overexpressed upon synaptic dysfunction and glial response, on methamphetamine-induced striatal dysfunction. The aim of this work was to provide an integrative characterization of the striatal changes in amino acids, monoamines and astroglia, as well as in the RAGE levels, and the associated motor activity profile of C57BL/6 adult mice, 72 h after a single-high dose of methamphetamine (30 mg/kg, i.p.). Our findings indicate, for the first time, that methamphetamine decreases striatal glutamine, glutamate and GABA levels, as well as glutamine/glutamate and GABA/glutamate ratios, while serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) levels remain unchanged. This methamphetamine regimen also produced dopaminergic terminal degeneration in the striatum, as evidenced by dopamine and tyrosine hydroxylase depletion. Consistently, methamphetamine decreased the locomotor activity of mice, in the open field test. In addition, increased levels of glutamine synthase and glial fibrillary acidic protein were observed. Nevertheless, methamphetamine failed to change RAGE levels. Our results show that acute methamphetamine intoxication induces pronounced changes in the striatal glutamatergic/GABAergic and dopaminergic homeostasis, along with astrocyte activation. These neurochemical and glial alterations are accompanied by impairment in locomotor activity.

Acute increase of the glutamate-glutamine cycling in discrete brain areas after administration of a single dose of amphetamine.

The glutamate–glutamine cycle between neurons and glia is tightly related to excitatory glutamatergic and inhibitory GABAergic regulation in brain. The role of this neuron–astrocyte cross‐talk on the neurotoxicity induced by amphetamines is not understood. Also, the impact of neurotoxic doses of amphetamines on the balance between glutamatergic and GABAergic circuits is largely unknown. The aim of this work was to assess the acute effect of a neurotoxic regimen of amphetamine (AMPH) on glutamine (GLN, an astrocytic marker) levels and on glutamine/glutamate (an index for glutamate–glutamine cycle) and GABA/glutamate ratios in rat brain. Sprague‐Dawley rats were sacrificed 4 and 24 h after a single‐dose regimen of AMPH (30 mg/kg, i.p.), and the caudate–putamen (CPu), frontal cortex (FC), and hippocampus (Hp) were dissected for analysis of glutamate (GLU), γ‐aminobutyric acid (GABA), and GLN. The total content of these amino acids was measured using a microbore HPLC electrochemical detector. Although AMPH did not change GLU levels, it increased both GLN content and GLN/GLU ratio (160–469%) at 4 h, but not at 24 h, in all regions after injection. Striatal GABA levels and GABA/GLU ratio were increased (46 and 100%, respectively) at 24 h. In hippocampus the GABA/GLU increase (60%) occurred as early as 4 h after treatment. To the contrary, AMPH exerted no effect in GABA/GLU balance in frontal cortex. These data strongly suggest that this neurotoxic AMPH regimen provoked an early increase in the glutamate–glutamine cycle between neurons and glia. This increase may ultimately lead to an upregulation of the inhibitory system as a compensatory response.

Dexamethasone Effect on Postoperative Pain and Tramadol Requirement after Thyroidectomy

Tramadol is a central-acting analgesic associated with nausea and vomiting. Clinical studies have demonstrated that glucocorticoids have analgesic and antiemetic effects when administered perioperatively. The aim of this study is to test the hypothesis that coadministration of tramadol and dexamethasone decreases both postoperative pain and tramadol requirement by patient-controlled analgesia (PCA). Forty female patients undergoing thyroidectomy under general anesthesia were enrolled in a double-blind randomized controlled study and allocated to receive dexamethasone 4 mg i.v. (dexamethasone group, n = 20) or saline (control group, n = 20). At 0, 1, 2, 4 and 22 h of PCA, tramadol consumption and pain were evaluated. Although pain (numerical rating scale 0–10) was significantly lower in the dexamethasone group compared to the control group (2.9 ± 1.4 vs. 3.8 ± 1.2, p = 0.02) at the beginning of PCA, tramadol demand was not significantly different. Although the results herein show a possible beneficial effect of a preoperative single low dose of dexamethasone on postoperative pain, the hypothesis that this corticosteroid decreases tramadol requirement is not supported.

Modulation of glucose-induced insulin secretion by cytosolic redox state in clonal beta-cells.

Nutrient stimulation of pancreatic beta-cells increases the cellular reduced pyridine nucleotide content, but the specific role of cytosolic redox state in glucose-induced insulin release (GIIR) remains undetermined. The role of cytosolic redox state has been assessed (as reflected by the lactate/pyruvate ratio) in nutrient- and non-nutrient-induced insulin release using a recently established glucose-sensitive clonal beta-cell line (BRIN-BD11). Long-term exposure to the NAD+ precursor vitamin nicotinic acid (NA, 100 microM) was used to promote a more oxidized state in the cytosol. Glucose (2-16 mM) evoked a dose-dependent rise in the cytosolic NADH/NAD+ ratio which was linearly related to the extent of GIIR. NA suppressed the glucose-induced rise in the NADH/NAD+ ratio and concomitantly reduced GIIR by 44%. It also inhibited, by 47%, the average glucose-induced rise in cytosolic free Ca2+ concentration ([Ca2+]i, assessed by fura-2 microfluorometry from single cells). The latter effect was not accounted for by a reduction in the activity of voltage-sensitive Ca2+ channels, inasmuch as both high K+- and tolbutamide-induced [Ca2+]i rises remained insensitive to NA exposure. NA did not affect insulin release evoked by any of the depolarizing agents, indicating that steps in the stimulus-secretion coupling cascade distal to Ca2+ influx are insensitive to changes in the cytosolic redox state. It is concluded that GIIR is partially controlled by the cytosolic redox state. Moreover, the impairment in GIIR, caused by a shift toward a more oxidized state in the cytosol, originates from an attenuated [Ca2+]i response. The latter is likely mediated by the influence of cytosolic redox state on specific metabolic pathways (NADH shuttle systems and/or the malonyl-CoA pathway), leading ultimately to enhancement of the activity of ATP-sensitive K+ channels.

Adaptation to Repeated Cocaine Administration in Rats

Abstract: Quantitative electroencephalogram (EEG) studies in cocaine‐dependent human patients show deficits in slow‐wave brain activity, reflected in diminished EEG power in the delta and theta frequency bands. In the present study, electrophysiological measures were monitored in 10 nonanesthetized, adult male Sprague‐Dawley rats via bipolar, epidural electrodes implanted over the somatosensory cortex. Control electrocorticograms (ECoG) were recorded twice within a two‐week interval to establish a baseline. Rats were subsequently injected daily with cocaine HCl at 15 mg/kg, i.p., for two weeks. The ECoG was recorded during a 1‐h session one day after the last injection. Total concentrations of dopamine (DA) and its metabolites were assayed in caudate nucleus (CN) and frontal cortex (FC) using HPLC/EC. Compared with controls, marked increases in DA concentrations were observed in both regions. The DA turnover decreased significantly. The power spectra, obtained by use of a fast Fourier transformation, revealed a significant decrease in slow‐wave delta frequency bands following repeated exposure to cocaine. These data are consistent with reported findings in humans that repeated exposures to cocaine result in a decrease in slow‐wave brain activity. Further studies are necessary to establish whether regional alterations in blood flow and metabolic activity may underlie such observations.