Overexpression of hyperpolarization-activated cyclic nucleotide-gated channels into the ventral tegmental area increases the rewarding effects of ethanol in UChB drinking rats.

BACKGROUND: A number of studies have shown that ethanol (EtOH) activates dopamine neurocircuitries and is self-administered into the ventral tegmental area (VTA) of the rat brain. In vitro and in silico studies have showed that hyperpolarization-activated cyclic nucleotide-gated (HCN) ionic channels on VTA dopamine neurons may constitute a molecular target of EtOH; however, there is no in vivo evidence supporting this assumption. METHODS: Wistar-derived University of Chile Drinking (UChB) rats were microinjected into the VTA with a lentiviral vector coding for rat HCN-2 ionic channel or a control vector. Four days after vector administration, daily voluntary EtOH intake was assessed for 30 days under a free-access paradigm to 5% EtOH and water. After EtOH consumption studies, the effect of HCN-2 overexpression was also assessed on EtOH-induced conditioned place preference (CPP); EtOH-induced locomotion, and EtOH-induced dopamine release in the nucleus accumbens (NAcc). RESULTS: Rats microinjected with the HCN-2 coding vector into the VTA showed (i) a ~2-fold increase in their voluntary EtOH intake compared to control animals, (ii) lentiviral-HCN-2-treated animals also showed an increased CPP to EtOH (~3-fold), (iii) a significant higher locomotor activity (~2-fold), and (iv) increased dopamine release in NAcc upon systemic administration of EtOH (~2-fold). CONCLUSIONS: Overexpression of HCN-2 ionic channel in the VTA of rats results in an increase in voluntary EtOH intake, EtOH-induced CPP, locomotor activity, and dopamine release in NAcc, suggesting that HCN levels in the VTA are relevant for the rewarding properties of EtOH.

Ethanol as a prodrug: brain metabolism of ethanol mediates its reinforcing effects.

BACKGROUND: While the molecular entity responsible for the rewarding effects of virtually all drugs of abuse is known, that for ethanol remains uncertain. Some lines of evidence suggest that the rewarding effects of alcohol are mediated not by ethanol per se but by acetaldehyde generated by catalase in the brain. However, the lack of specific inhibitors of catalase has not allowed strong conclusions to be drawn about its role on the rewarding properties of ethanol. The present studies determined the effect on voluntary alcohol consumption of two gene vectors, one designed to inhibit catalase synthesis and one designed to synthesize alcohol dehydrogenase (ADH), to respectively inhibit or increase brain acetaldehyde synthesis. METHODS: The lentiviral vectors, which incorporate the genes they carry into the cell genome, were (i) one encoding a shRNA anticatalase synthesis and (ii) one encoding alcohol dehydrogenase (rADH1). These were stereotaxically microinjected into the brain ventral tegmental area (VTA) of Wistar-derived rats bred for generations for their high alcohol preference (UChB), which were allowed access to an ethanol solution and water. RESULTS: Microinjection into the VTA of the lentiviral vector encoding the anticatalase shRNA virtually abolished (-94% p < 0.001) the voluntary consumption of alcohol by the rats. Conversely, injection into the VTA of the lentiviral vector coding for ADH greatly stimulated (2 to 3 fold p < 0.001) their voluntary ethanol consumption. CONCLUSIONS: The study strongly suggests that to generate reward and reinforcement, ethanol must be metabolized into acetaldehyde in the brain. Data suggest novel targets for interventions aimed at reducing chronic alcohol intake.

Nicotinamide prevents the long-term effects of perinatal asphyxia on apoptosis, non-spatial working memory and anxiety in rats.

There is no established treatment for the long-term effects produced by perinatal asphyxia. Thus, we investigated the neuroprotection provided by nicotinamide against the effects elicited by perinatal asphyxia on hippocampus and behaviour observed at 30-90 days of age. Asphyxia was induced by immersing foetuses-containing uterine horns, removed from ready-to-deliver rats into a water bath at 37 degrees C for 20 min. Caesarean-delivered siblings were used as controls. Saline or nicotinamide (0.8 mmol/kg, i.p.) was administered to control and asphyxia-exposed animals 24, 48, and 72 h after birth. The animals were examined for morphological changes in hippocampus, focusing on delayed cell death and mossy fibre sprouting, and behaviour, focusing on cognitive behaviour and anxiety. At the age of 30-45 days, asphyxia-exposed rats displayed (1) increased apoptosis, assessed in whole hippocampus by nuclear Hoechst staining, and (2) increased mossy fibre sprouting, restricted to the stratum oriens of dorsal hippocampus, assessed by Timm's staining. Rats from the same cohorts displayed (3) deficits in non-spatial working memory, assessed by a novel object recognition task, and (4) increased anxiety, assessed by an elevated plus-maze test when examined at the age of 90 days. Nicotinamide prevented the effects elicited by perinatal asphyxia on apoptosis, working memory, and anxiety.

Rat strain influences the use of egocentric learning strategies mediated by neostriatum.

Rats use place (allocentric) or stimulus-response (egocentric) learning strategies for foraging under ethological and/or experimental conditions, proposed to be conveyed by hippocampus or neostriatum, respectively. We investigated here the effect of a reversible blockade of neostriatum on learning strategies assessed by a cross maze paradigm, comparing A x C (phenotypically similar to wild rats) versus Long-Evans rat strains. The rats were trained to reach a consistently baited-arm (west arm), starting from the same arm (south arm). The learning strategy was evaluated at days 11 and 19, when test trials were performed placing the rat in a start-box at the arm (north arm) opposite to that when starting the training, following a saline or lidocaine injection into the neostriatum. Rats entering to the baited-west arm were considered to be place learners and those entering to the unbaited-east arm were response learners. It was found that Long-Evans rats injected with saline were place learners on day 11 and response learners on day 19, but were place learners on both days when injected with lidocaine. A x C rats injected with saline were response learners on days 11 and 19, and place learners on both days when injected with lidocaine. Thus, rat strain influences the memory strategy for solving a cross maze paradigm. Long-Evans, but not A x C rats, shift from place (allocentric) to response (egocentric) learning along the training. When neostriatum was blocked by lidocaine, both rat strains elicited a place learning strategy only.

Ethanol induces stronger dopamine release in nucleus accumbens (shell) of alcohol-preferring (bibulous) than in alcohol-avoiding (abstainer) rats.

Several studies on the differences between ethanol-preferring versus non-preferring rat lines suggest an innate deficit in the mesolimbic dopaminergic system as an underlying factor for ethanol volition. Rats would try to overcome such deficit by engaging in a drug-seeking behaviour, when available, to drink an ethanol solution over water. Thus, in the present study we compared the effect of a single dose of ethanol (1 g/kg, i.p.) on the extracellular levels of monoamines measured by microdialysis in the shell of nucleus accumbens of University of Chile bibulous (UChB) and University of Chile Abstainer (UChA) rats, bred for 79 and 88 generations to prefer or reject ethanol, respectively. It is reported that under basal conditions extracellular dopamine levels are lower in the bibulous than in the abstainer rats, while ethanol induced a 2-fold greater increase of dopamine release in bibulous than in abstainer rats. The greater effect of ethanol in bibulous rats was not associated to differences in blood ethanol levels, since the concentration and elimination of ethanol were virtually identical in both rat lines, indicating that bibulous rats are more sensitive to the stimulation of dopamine release by ethanol than abstainer rats. No differences were observed in 5-hydroxytryptamine or metabolites measured simultaneously under basal or ethanol-stimulating conditions in bibulous and abstainer rats. Overall, the present results suggest that a low dopaminergic tone and a strong mesolimbic dopamine response to ethanol are concerted neurochemical features associated to an ethanol-seeking behaviour in rats.

Acute perinatal asphyxia impairs non-spatial memory and alters motor coordination in adult male rats.

A large body of clinical evidence suggests a possible association between perinatal asphyxia and the onset of early, as well as long-term, neurological and psychiatric disorders including cognitive deficits. The present study investigated cognitive and motor function modifications in a well characterized and clinically relevant experimental rat model of human perinatal asphyxia. The results reported here show that adult rats exposed to a single (20 min) asphyctic episode at delivery displayed: (a) a deficit in non-spatial memory, assessed in a novel object recognition task; (b) an impaired motor coordination, measured by the rotarod test. On the other hand, gross motor activity and spatial memory, evaluated in both the Y maze and the Barnes maze, were not affected by perinatal asphyxia. The results of this study provide further insights into the long-term effects of perinatal asphyxia on neurobehavioural functions.

Targeting Sentinel Proteins and Extrasynaptic Glutamate Receptors: a Therapeutic Strategy for Preventing the Effects Elicited by Perinatal Asphyxia?

Perinatal asphyxia (PA) is a relevant cause of death at the time of labour, and when survival is stabilised, associated with short- and long-term developmental disabilities, requiring inordinate care by health systems and families. Its prevalence is high (1 to 10/1000 live births) worldwide. At present, there are few therapeutic options, apart from hypothermia, that regrettably provides only limited protection if applied shortly after the insult.PA implies a primary and a secondary insult. The primary insult relates to the lack of oxygen, and the secondary one to the oxidative stress triggered by re-oxygenation, formation of reactive oxygen (ROS) and reactive nitrogen (RNS) species, and overactivation of glutamate receptors and mitochondrial deficiencies. PA induces overactivation of a number of sentinel proteins, including hypoxia-induced factor-1α (HIF-1α) and the genome-protecting poly(ADP-ribose) polymerase-1 (PARP-1). Upon activation, PARP-1 consumes high amounts of ATP at a time when this metabolite is scarce, worsening in turn the energy crisis elicited by asphyxia. The energy crisis also impairs ATP-dependent transport, including glutamate re-uptake by astroglia. Nicotinamide, a PARP-1 inhibitor, protects against the metabolic cascade elicited by the primary stage, avoiding NAD+ exhaustion and the energetic crisis. Upon re-oxygenation, however, oxidative stress leads to nuclear translocation of the NF-κB subunit p65, overexpression of the pro-inflammatory cytokines IL-1ß and TNF-α, and glutamate-excitotoxicity, due to impairment of glial-glutamate transport, extracellular glutamate overflow, and overactivation of NMDA receptors, mainly of the extrasynaptic type. This leads to calcium influx, mitochondrial impairment, and inactivation of antioxidant enzymes, increasing further the activity of pro-oxidant enzymes, thereby making the surviving neonate vulnerable to recurrent metabolic insults whenever oxidative stress is involved. Here, we discuss evidence showing that (i) inhibition of PARP-1 overactivation by nicotinamide and (ii) inhibition of extrasynaptic NMDA receptor overactivation by memantine can prevent the short- and long-term consequences of PA. These hypotheses have been evaluated in a rat preclinical model of PA, aiming to identify the metabolic cascades responsible for the long-term consequences induced by the insult, also assessing postnatal vulnerability to recurrent oxidative insults. Thus, we present and discuss evidence demonstrating that PA induces long-term changes in metabolic pathways related to energy and oxidative stress, priming vulnerability of cells with both the neuronal and the glial phenotype. The effects induced by PA are region dependent, the substantia nigra being particularly prone to cell death. The issue of short- and long-term consequences of PA provides a framework for addressing a fundamental issue referred to plasticity of the CNS, since the perinatal insult triggers a domino-like sequence of events making the developing individual vulnerable to recurrent adverse conditions, decreasing his/her coping repertoire because of a relevant insult occurring at birth.

Dopamine release in the nucleus accumbens (shell) of two lines of rats selectively bred to prefer or avoid ethanol.

Lower tissue levels of dopamine and 5-hydroxytryptamine (5-HT) have been found in the nucleus accumbens of alcohol-naïve rats selectively bred to prefer ethanol than in rats bred to avoid it. These findings have led to the hypothesis that differences in the dopamine and 5-HT tone may be linked to ethanol preference. In the present study we used the in vivo microdialysis technique to determine the actual extracellular levels of dopamine, its metabolites 3,4-dihydroxyphenyl acetaldehyde (DOPALD), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 5-HT and 5-hydroxyindolacetic acid (5-HIAA) in the shell of nucleus accumbens of rat lines selectively bred as either high-ethanol (UChB) or low-ethanol (UChA) drinkers. Basal extracellular levels of dopamine, DOPALD, DOPAC and HVA were lower in the shell of nucleus accumbens of ethanol-naïve UChB than in UChA rats. In agreement, when perfused with 100 microM d-amphetamine or 100 mM KCl lower dopamine increases were observed in nucleus accumbens of UChB rats compared to UChA rats, indicating lower cytosolic (d-amphetamine releasable) and vesicular (KCl releasable) dopamine pools in UChB animals. Since the experiments were performed in ethanol-naïve rats, the present results suggest an innate deficiency in the mesolimbic dopamine system of UChB rats. There were no line differences in basal, d-amphetamine or KCl stimulated 5-HT levels. Thus, the present findings support a role of dopamine, but not of 5-HT, as predictor of ethanol preference in UChB rats. Overall, data obtained are in agreement with previous reports in other rat lines showing that lower dopamine levels and its metabolites are associated with a genetic predisposition to ethanol preference.

Exploring neurocircuitries of the basal ganglia by intracerebral administration of selective neurotoxins.

The detailed anatomy of the monoamine pathways of the rat, first described by the students of Nils Ake Hillarp in Sweden, provided the basis for a neurocircuitry targeted pharmacology, leading to important therapeutic breakthroughs. Progress was achieved by the introduction of accurate lesion techniques based on selective neurotoxins. Systematic intracerebral injections of 6-hydroxydopamine let Urban Ungerstedt at the Karolinska Institutet, Stockholm, Sweden, to propose the first stereotaxic mapping of the monoamine pathways in the rat brain; and the 'Rotational Behaviour', as a classical model for screening drugs useful for alleviating Parkinson's disease and other neuropathologies. The direction of the rotational behaviour induced by drugs administrated to unilaterally 6-hydroxydopamine-lesioned rats reveals their mechanism of action at dopamine synapses, as demonstrated when rotational behaviour was combined with microdialysis. The model was useful for proposing a role for dopamine receptors in the gating of the flow of information integrated and/or modulated by the basal ganglia, through different efferent pathways; notably the striatopallidal system, via D(2) receptors, and the striatonigral system, via D(1) receptors. The role of other dopamine receptor subtypes on rotational behaviour has not yet been clarified.

Neurochemical and behavioural characterisation of alkoxyamphetamine derivatives in rats.

The clinical utility of amphetamine and amphetamine analogues has been jeopardized by a number of side effects and toxicity, partly due to complex mechanisms of action. While some of the analogues have been individually characterised, there is still need for comparative studies, in particularly on their efficacy to release dopamine and 5-hydroxytryptamine, further enlightening some of the synaptic mechanisms conveying their actions. Thus, we have compared four alkoxyamphetamine derivatives, i.e., p-methoxyamphetamine; p-methoxymethamphetamine; methylenedioxyamphetamine, methylenedioxymethamphetamine, using methamphetamine, and D-amphetamine, as reference substances, on rotational behaviour and releasing mechanisms studied with in vivo microdialysis in rats. All alkoxylated-derivatives produced a long-lasting rotational behaviour at 10 mg/kg s.c., but the reference substances produced a strong rotation already at 2 mg/kg s.c. in 6-hydroxydopamine-lesioned rats. At the concentration of 100 micromolar, the alkoxylated-derivatives were equipotent to evoke dopamine and 5-hydroxytryptamine release in rat neostriatum, while D-amphetamine and methamphetamine were more efficient on dopamine release. Pre-treatment with methamphetamine or the alkoxylated-derivatives produced a remarkable decrease of the effect of K+ -depolarisation on both dopamine and 5-hydroxytryptamine release. The insertion of a methoxy or a methylenedioxy group on the benzene ring of D-amphetamine or methamphetamine, or N-methylation of the D-amphetamine molecule alters the selectivity of the compounds. The efficacy of the alkoxylated-derivatives on dopamine and 5-hydroxytryptamine release was similar, but stimulated less dopamine release and produced less rotational behaviour than D-amphetamine and methamphetamine. The lower efficacy of K+ -depolarisation following pre-treatments with the derivatives suggests an impairment of releasable monoamine stores. The present observations can enlighten the mechanisms of action of drugs showing a high risk for abuse among young populations.

Effect of perinatal asphyxia on tuberomammillary nucleus neuronal density and object recognition memory: A possible role for histamine?

Perinatal asphyxia (PA) is associated with long-term neuronal damage and cognitive deficits in adulthood, such as learning and memory disabilities. After PA, specific brain regions are compromised, including neocortex, hippocampus, basal ganglia, and ascending neuromodulatory pathways, such as dopamine system, explaining some of the cognitive disabilities. We hypothesize that other neuromodulatory systems, such as histamine system from the tuberomammillary nucleus (TMN), which widely project to telencephalon, shown to be relevant for learning and memory, may be compromised by PA. We investigated here the effect of PA on (i) Density and neuronal activity of TMN neurons by double immunoreactivity for adenosine deaminase (ADA) and c-Fos, as marker for histaminergic neurons and neuronal activity respectively. (ii) Expression of the histamine-synthesizing enzyme, histidine decarboxylase (HDC) by western blot and (iii) thioperamide an H3 histamine receptor antagonist, on an object recognition memory task. Asphyxia-exposed rats showed a decrease of ADA density and c-Fos activity in TMN, and decrease of HDC expression in hypothalamus. Asphyxia-exposed rats also showed a low performance in object recognition memory compared to caesarean-delivered controls, which was reverted in a dose-dependent manner by the H3 antagonist thioperamide (5-10mg/kg, i.p.). The present results show that the histaminergic neuronal system of the TMN is involved in the long-term effects induced by PA, affecting learning and memory.

Quantitative genetics of bioenergetics and growth-related traits in the wild mammal, Phyllotis darwini.

We studied the potential for response to selection in typical physiological-thermoregulatory traits of mammals such as maximum metabolic rate (MMR), nonshivering thermogenesis (NST) and basal metabolic rate (BMR) on cold-acclimated animals. We used an animal model approach to estimate both narrow-sense heritabilities (h2) and genetic correlations between physiological and growth-related traits. Univariate analyses showed that MMR presented high, significant heritability (h2 = 0.69 +/- 0.35, asymptotic standard error), suggesting the potential for microevolution in this variable. However, NST and BMR presented low, nonsignificant h2, and NST showed large maternal/common environmental/nonadditive effects (c2 = 0.34 +/- 0.17). Heritabilities were large and significant (h2 > 0.5) for all growth-related traits (birth mass, growth rate, weaning mass). The only significant genetic correlations we found between a physiological trait and a growth-related trait was between NST and birth mass (r = -0.74; P < 0.05). Overall, these results suggest that additive genetic variance is present in several bioenergetic traits, and that genetic correlations could be present between those different kinds of traits.

The quantitative genetics of sustained energy budget in a wild mouse.

We explored how morphological and physiological traits associated with energy expenditure over long periods of cold exposure would be integrated in a potential response to natural selection in a wild mammal, Phyllotis danwini. In particular, we studied sustained energy expenditure (SusMR), the rate of expenditure fueled by concurrent energy intake, basal metabolic rate (BMR), and sustained metabolic scope (SusMS = SusMR/BMR), a measure of the reserve for sustained work. We included the masses of different central processing organs as an underlying factor that could have a mechanistic link with whole animal traits. Only the liver had heritability statistically different from zero (0.73). Physiological and morphological traits had high levels of specific environmental variance (average 70%) and postnatal common environmental variance (average 30%) which could explain the low heritabilities estimates. Our results, (1) are in accordance with previous studies in mammals that report low heritabilities for metabolic traits (SusMR, BMR, SusMS), (2) but not completely with previous ones that report high heritabilities for morphological traits (masses of central organs), and (3) provide important evidence of the relevance of postnatal common environmental variance to sustained energy expenditure.

Effects of the DT-diaphorase inhibitor dicumarol on striatal monoamine levels in L-DOPA and L-deprenyl pre-treated rats.

It has been proposed that DT-diaphorase plays a strategic role as a neuroprotective enzyme for monoamine neurons, perhaps together with monoamine oxidase (MAO). Thus, we investigated the long-term effects produced by DT-diaphorase inhibition with dicumarol injected unilaterally into the medial forebrain bundle (MFB) on monoamine and metabolite levels, alone, or following dopamine loading with 3,4-dihydroxyphenyl-L-alanine (L-DOPA) or MAO inhibition with L-deprenyl. Monoamine levels were assayed in aliquots from tissue samples from right and left striatum, including both dorsal and ventral regions. Dicumarol alone produced increases in 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA), but not in dopamine and metabolite levels when assayed two weeks later. However, following preloading with L-DOPA (3 x 25 mg/kg s.c. 7, 4 and 1 h before surgery), a long-lasting bilateral increase in dopamine and metabolite levels was observed after dicumarol. No effect was observed on dopamine, 5-HT and metabolite levels after L-deprenyl (3 x 10 mg/kg, s.c.) alone, but the levels were unilaterally increased when L-deprenyl was followed by dicumarol. The same result was produced when both L-deprenyl and dicumarol were injected simultaneously into the same brain region. In conclusion, the present study shows that intracerebral inhibition of DT-diaphorase produces long-term changes in 5-HT, but also in dopamine metabolism when DT-diaphorase inhibition is combined with MAO inhibition by systemic or intracerebral treatment with L-deprenyl. It is suggested that both MAO and DT-diaphorase have to be inhibited for inducing long-term changes in monoamine metabolism. Thus, DT-diaphorase is an enzyme to be taken into account when L-DOPA is used to treat Parkinson's disease, or when an MAO-inhibitor is used to treat depression.

Analgesic and behavioral effects of amphetamine enantiomers, p-methoxyamphetamine and n-alkyl-p-methoxyamphetamine derivatives.

The analgesic effects of (+)- and (-)-amphetamine (AMPH), (+/-)-p-methoxyamphetamine (MA), (+/-)-N-methyl-p-methoxyamphetamine (MMA) and (+/-)-N-ethyl-p-methoxyamphetamine (EMA) were compared using two different algesimetric tests in rats. In the formalin test, (+)-AMPH elicited significant antinociception at doses of 0.2, 2 and 8 mg/kg (i.p.); (-)-AMPH was active at 2 and 8 mg/kg, but not at 0.2 mg/kg; MA elicited very potent and long-lasting antinociception; MMA was less active than MA; EMA showed significant effects only at doses of 2 and 8 mg/kg. In the C-fiber evoked nociceptive reflex assay, i.v. (+)- and (-)-AMPH were ineffective, but the methoxy derivatives showed a similar pattern of action combining inhibitory and excitatory actions. To clarify apparent discrepancies between both algesimetric tests, some behavioral motor performance tests were carried out. These tests confirm the motor stimulatory properties of (+)-AMPH, not shared by the methoxylated amphetamine derivatives. The three methoxy derivatives elicited some stereotypies related to dopaminergic activation such as grooming behavior. (+)-AMPH was also the only drug to increase the acquisition of CARs while MA and EMA were without effect. Avoidance conditioning was seriously impaired in rats injected with MMA. This conditioned behavior can be related to the significant decrease of spontaneous motor activity observed with this drug. In conclusion, the introduction of a para-methoxy group strongly increases the analgesic effects of amphetamine without its stimulatory behavioral effects. The introduction of N-alkyl substituents decreases the analgesic potency of MA.

Dynamic thermal balance in the leaf-eared mouse: the interplay among ambient temperature, body size, and behavior.

Endotherms maintain constant body temperature through physiological and behavioral adjustments. Behavioral thermoregulation is an important factor influencing energy balance. We exposed the leaf-eared mouse, Phyllotis darwini, to temperatures corresponding to its natural thermal range and studied two forms of behavioral thermoregulation: diminishing surface to volume ratio by huddling and heat dissipation by increasing physical contact with the substrate (flattening). We predicted that at low ambient temperatures (T(a)) huddling would be used as a heat conservation mechanism and at high T(a) flattening would be used for heat loss. We simultaneously measured oxygen consumption (VO2) and flattening, in response to three independent factors: huddling, T(a), and body mass. Each experiment was a 6-h VO2 trial where five virgin females were measured at constant T(a). We performed this protocol for two body mass groups, small (ca. 40 g) and large (ca. 70 g), in a metabolic chamber. Treatments were groups with and without the ability to huddle at five different T(a), ranging from 5 degrees to 35 degrees C. A significant interaction between all three factors was found. Huddling and flattening were used as strategies for conserving or dissipating heat, respectively, and the shift between both strategies occurred at the lower limit of thermoneutrality. At T(a) below thermoneutrality, huddling was a more effective way of reducing metabolic requirements and was more efficient (H(E)) in small individuals than large individuals. So, by huddling, small individuals save more energy. At high T(a), flattening appeared to be an equally useful mechanism for heat loss, for both large and small animals.

Salsolinol, free of isosalsolinol, exerts ethanol-like motivational/sensitization effects leading to increases in ethanol intake.

Salsolinol is formed non-enzymatically when ethanol-derived acetaldehyde binds to dopamine, yielding 2 distinct products, i.e., salsolinol and isosalsolinol. Early animal studies, revealing that salsolinol promotes alcohol consumption and recent evidence that animals will readily self-administer salsolinol into the posterior ventral tegmental area (p-VTA) together with the finding that salsolinol is able to induce conditioned place preference and to increase locomotor activity, have outlined a role of salsolinol in the behavioral and neurobiological actions of ethanol. Until recently, the only commercially available salsolinol was a mixture containing 85% salsolinol and 10-15% isosalsolinol. The possibility thus exists that either salsolinol or isosalsolinol explains the reinforcing properties of ethanol. We report here that a newly available salsolinol is free of isosalsolinol. Thus, salsolinol, free of isosalsolinol, was injected intracerebrally (30 pmol/0.2 µL, into the ventral tegmental area [VTA]) or intraperitoneally (i.p.) (10 mg/kg) to naïve rats bred as alcohol drinkers to study salsolinol's motivational effects and its role on voluntary ethanol intake. Salsolinol produced conditioned place preference and increased locomotor activity, whether injected intra-VTA or intraperitoneally. Following systemic (i.p.) administration of 10 mg/kg salsolinol, this molecule was detected in vivo by microdialysis of neostriatum, reaching an estimated concentration of 100 nM in the dialyzate. These results indicate that systemically administered salsolinol is able to cross the blood-brain barrier (BBB). Repeated administration of salsolinol sensitized rats to the locomotor activity and led to increases in voluntary ethanol consumption, which was prevented by intra-VTA pretreatment with naltrexone.

Perinatal asphyxia: CNS development and deficits with delayed onset.

Perinatal asphyxia constitutes a prototype of obstetric complications occurring when pulmonary oxygenation is delayed or interrupted. The primary insult relates to the duration of the period lacking oxygenation, leading to death if not re-established. Re-oxygenation leads to a secondary insult, related to a cascade of biochemical events required for restoring proper function. Perinatal asphyxia interferes with neonatal development, resulting in long-term deficits associated to mental and neurological diseases with delayed clinical onset, by mechanisms not yet clarified. In the experimental scenario, the effects observed long after perinatal asphyxia have been explained by overexpression of sentinel proteins, such as poly(ADP-ribose) polymerase-1 (PARP-1), competing for NAD(+) during re-oxygenation, leading to the idea that sentinel protein inhibition constitutes a suitable therapeutic strategy. Asphyxia induces transcriptional activation of pro-inflammatory factors, in tandem with PARP-1 overactivation, and pharmacologically induced PARP-1 inhibition also down-regulates the expression of proinflammatory cytokines. Nicotinamide has been proposed as a suitable PARP-1 inhibitor. Its effect has been studied in an experimental model of global hypoxia in rats. In that model, the insult is induced by immersing rat fetus into a water bath for various periods of time. Following asphyxia, the pups are delivered, treated, and nursed by surrogate dams, pending further experiments. Nicotinamide rapidly distributes into the brain following systemic administration, reaching steady state concentrations sufficient to inhibit PARP-1 activity for several hours, preventing several of the long-term consequences of perinatal asphyxia, supporting the idea that nicotinamide constitutes a lead for exploring compounds with similar or better pharmacological profiles.

Pathophysiology of perinatal asphyxia: can we predict and improve individual outcomes?

Perinatal asphyxia occurs still with great incidence whenever delivery is prolonged, despite improvements in perinatal care. After asphyxia, infants can suffer from short- to long-term neurological sequelae, their severity depend upon the extent of the insult, the metabolic imbalance during the re-oxygenation period and the developmental state of the affected regions. Significant progresses in understanding of perinatal asphyxia pathophysiology have achieved. However, predictive diagnostics and personalised therapeutic interventions are still under initial development. Now the emphasis is on early non-invasive diagnosis approach, as well as, in identifying new therapeutic targets to improve individual outcomes. In this review we discuss (i) specific biomarkers for early prediction of perinatal asphyxia outcome; (ii) short and long term sequelae; (iii) neurocircuitries involved; (iv) molecular pathways; (v) neuroinflammation systems; (vi) endogenous brain rescue systems, including activation of sentinel proteins and neurogenesis; and (vii) therapeutic targets for preventing or mitigating the effects produced by asphyxia.

Perinatal asphyxia: current status and approaches towards neuroprotective strategies, with focus on sentinel proteins.

Delivery is a stressful and risky event menacing the newborn. The mother-dependent respiration has to be replaced by autonomous pulmonary breathing immediately after delivery. If delayed, it may lead to deficient oxygen supply compromising survival and development of the central nervous system. Lack of oxygen availability gives rise to depletion of NAD(+) tissue stores, decrease of ATP formation, weakening of the electron transport pump and anaerobic metabolism and acidosis, leading necessarily to death if oxygenation is not promptly re-established. Re-oxygenation triggers a cascade of compensatory biochemical events to restore function, which may be accompanied by improper homeostasis and oxidative stress. Consequences may be incomplete recovery, or excess reactions that worsen the biological outcome by disturbed metabolism and/or imbalance produced by over-expression of alternative metabolic pathways. Perinatal asphyxia has been associated with severe neurological and psychiatric sequelae with delayed clinical onset. No specific treatments have yet been established. In the clinical setting, after resuscitation of an infant with birth asphyxia, the emphasis is on supportive therapy. Several interventions have been proposed to attenuate secondary neuronal injuries elicited by asphyxia, including hypothermia. Although promising, the clinical efficacy of hypothermia has not been fully demonstrated. It is evident that new approaches are warranted. The purpose of this review is to discuss the concept of sentinel proteins as targets for neuroprotection. Several sentinel proteins have been described to protect the integrity of the genome (e.g. PARP-1; XRCC1; DNA ligase IIIα; DNA polymerase ß, ERCC2, DNA-dependent protein kinases). They act by eliciting metabolic cascades leading to (i) activation of cell survival and neurotrophic pathways; (ii) early and delayed programmed cell death, and (iii) promotion of cell proliferation, differentiation, neuritogenesis and synaptogenesis. It is proposed that sentinel proteins can be used as markers for characterising long-term effects of perinatal asphyxia, and as targets for novel therapeutic development and innovative strategies for neonatal care.