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El rápido desarrollo de las neurotecnologías y la inteligencia artificial en el siglo XXI plantea nuevos desafíos éticos y preguntas acerca de las posibles amenazas que representaría el uso incorrecto o malintencionado de estas técnicas. Surge así el dilema del control: ¿debemos regular la neurotecnología antes de que se consolide, arriesgándonos así a frenar su desarrollo, o debemos esperar a que se afiance para evaluar entonces su impacto ético, aun a riesgo de que pueda resultar demasiado tarde? En apoyo de una respuesta dirigida hacia la regulación temprana, el presente trabajo se centra específicamente en el posible impacto de la neurotecnología para el neuroderecho a la identidad personal. Basaremos nuestro análisis en dos ejemplos: un estudio de caso y una situación hipotética. El estudio de caso tratará sobre los efectos del compuesto activo de la marihuana, el tetrahidrocannabinol (THC). La situación hipotética consistirá en la posibilidad futura de que se cumpla el escenario conocido como “cerebro en una cubeta”.

Este es el artículo que se ha publicado de forma definitiva en: https://pubs.rsc.org/en/content/articlepdf/2019/fo/c8fo02550a Obesity is a medical and sociological problem of great importance due to the high percentage of people affected and the important health consequences that it involves. Most cases of obesity are related to an inadequate diet, rich in fats, which could lead to changes in the patient’s oxygenic metabolism. That is why this study has been proposed to evaluate how some aspects of oxygenic metabolism are affected in a nutritional experimental model, with a controlled hyperlipidic liquid diet based on olive oil, and the effect of the antioxidant vitamin C on these conditions. Wistar rats were divided into four groups which received a control and hyperlipidic liquid diet for 30 days, with or without a vitamin C supplement (CO, COC, HO and HOC). First of all the body and fat tissue development was measured in the four groups. Our results showed that the excessive intake of nutritional and healthy fat such as olive oil did not prevent the appearance of obesity and the supplementation with vitamin C did not have a protective effect on body and fat development. The study of the antioxidant enzymes superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) in total liver, liver cytosol, abdominal white fat, brown fat and blood cells showed that vitamin C could have different selectivities and affinities for different enzymes and compartments/tissues of the body. Finally, the effect of vitamin C on various metabolic parameters (glucose, pyruvate, lactate, LDH, ATP, acetoacetate and beta-hydroxybutyrate) provided positive protection against oxidative stress especially under hyperlipidic conditions. All things considered, the present study concludes that vitamin C treatment could protect Wistar rats from the oxidative stress impairment induced by obesity generated by an excessive intake of fats.

Este artículo se encuentra disponible en la siguiente URL: https://www.frontiersin.org/articles/10.3389/fphys.2020.00463/full Myopia is one of the commonest eye pathologies that could affect 2.56 billion people by 2020. Today high myopia is a leading cause of blindness worldwide due to associated ocular illness. Nevertheless, the cellular bases for these diseases to develop are unclear in many areas. We conducted a prospective study of oxidative stress and growth factors in human myopic and non myopic eyes in an attempt to increase our understanding of the underlying physiopathological conditions to adequately early diagnose, prevent and treat the retina problem that derives from myopia. Aqueous humor samples were obtained from 41 patients being operated for cataracts in our hospital. Axial length, refractive status and complete ophthalmologic examination were recorded. The VEGF and HGF levels were determined by an ELISA kit. Total antioxidant capacity and total nitrites/nitrate levels were established with a lab kit. We show for the first time an increase in the total nitrite levels in high myopia. We also propose for the first time the concurrence of three factors: myopia, oxidative stress, and oxidative stress together with growth factors in the same group of patients. In this way, it would not be accurate to envision high myopia as a type of normal myopia, but one with more diopters or longer axial length.

Este artículo se encuentra disponible en la página web de la revista en la siguiente URL: https://www.frontiersin.org/articles/10.3389/fphys.2018.01008/full Retinitis pigmentosa (RP) is one of the most common clinical subtypes of retinal degeneration (RD), and it is a neurodegenerative disease that could cause complete blindness in humans because it ultimately affects the photoreceptors viability. RP afflicts an estimated 1.5 million patients worldwide. The retina is highly susceptible to oxidative stress which can impair mitochondrial function. Many retina pathologies, such as diabetic retinopathy and secondary cone photoreceptor death in RP, have been related directly or indirectly with mitochondrial dysfunction. The possible role of autophagy in retina and cell differentiation is described and also the implications of autophagy dysregulation in RP. The present review shows the crucial role of autophagy in maintaining the retina homeostasis and possible therapeutic approaches for the treatment of RP.

The intake of antioxidants has increased in the last years in order to treat some pathologies associated with oxidative stress such as cancer, diabetes mellitus, atherosclerosis, myocardial infarction, acute pancreatitis, Parkinson's and Alzheimer's disease among others. In colorectal cancer, a widely antineoplastic drug used is the fluorinated uracil molecule 5-Fluorouracile (5-FU). The aim of this study is to assess the antioxidant capacity observed by the inhibition of lipid peroxidation by 5-FU and other fluorinated uracil derivates: 6-[Difluoro(phenyl)methyl]-3-(2,4,6-trifluorophenyl)pyrimidine-2,4(1H,3H)-dione (S-81), 3-[2-Chloroethyl]-6-[difluoro(phenyl)methyl]pyrimidine-2,4(1H,3H)-dione (S-86), 6-[Difluoro(phenyl)methyl]-3-(4-trifluoromethoxyphenyl)pyrimidine-2,4(1H,3H)-dione (S-123). The results showed a significant decrease in MDA production of 58.12% in S-86, 44.61% in S-123, 24.11% in 5-FU and 10.83% in S-81 in the sample with highest concentration (10µM). 5-FU also showed a Total Antioxidant Capacity of 0.68 Trolox Equivalent Antioxidant Capacity.

Este artículo se encuentra disponible en la siguiente URL: https://www.frontiersin.org/articles/10.3389/fphys.2017.00022/full Alcohol is a legal drug present in several drinks commonly used worldwide (chemically known as ethyl alcohol or ethanol). Alcohol consumption is associated with several disease conditions, ranging from mental disorders to organic alterations. One of the most deleterious effects of ethanol metabolism is related to oxidative stress. This promotes cellular alterations associated with inflammatory processes that eventually lead to cell death or cell cycle arrest, among others. Alcohol intake leads to bone destruction and modifies the expression of interleukins, metalloproteinases and other pro-inflammatory signals involving GSKβ, Rho, and ERK pathways. Orthodontic treatment implicates mechanical forces on teeth. Interestingly, the extra- and intra-cellular responses of periodontal cells to mechanical movement show a suggestive similarity with the effects induced by ethanol metabolism on bone and other cell types. Several clinical traits such as age, presence of systemic diseases or pharmacological treatments, are taken into account when planning orthodontic treatments. However, little is known about the potential role of the oxidative conditions induced by ethanol intake as a possible setback for orthodontic treatment in adults.

Cannabis sativa is a plant that produces, among other cannabinoids, the psychoactive Δ-9-tetrahydrocannabinol (THC), whose adverse health effects have been recently reviewed. Repeated doses of THC lead to the development of tolerance to its own effects and the effects of other cannabinoids, and affect the same reward systems as alcohol, cocaine, and opioids. Enhanced brain oxidative stress usually correlates with cognitive impairment and a higher risk of development of neurodegenerative diseases, which has been repeatedly reported for many conditions. THC-induced cognitive effects are aggravated by the combined use of alcohol. Some research suggests similar mechanistic aspects of THC and alcohol on brain mitochondrial oxidative damage. Taking THC and ethanol together represents a greater risk for cognitive and attention impairment than taking either drug separately. While THC and ethanol exposure elevates reactive oxygen species in many tissues including the brain, further research is needed to establish a deleterious role of oxidative stress in the nervous system subjected to both compounds.

Molecular topology can be considered an application of graph theory in which the molecular structure is characterized through a set of graph-theoretical descriptors called topological indices. Molecular topology has found applications in many different fields, particularly in biology, chemistry, and pharmacology. The first topological index was introduced by H. Wiener in 1947 [1]. Although its very first application was the prediction of the boiling points of the alkanes, the Wiener index has demonstrated since then a predictive capability far beyond that. Along with the Wiener index, in this paper we focus on a few pioneering topological indices, just to illustrate the connection between physicochemical properties and molecular connectivity.

Molecular topology is an application of graph theory and statistics in fields like chemistry, biology, and pharmacology, in which the molecular structure matters. Its scope is the topological characterization of molecules by means of numerical invariants, called topological indices, which are the main ingredients of the molecular topological models. These are statistical models that are instrumental in the discovery of new applications of naturally occurring molecules, as well as in the design of synthetic molecules with specific chemical, biological, or pharmacological properties. In this review, we focus on pharmacology, which is a novel field of application of molecular topology. Besides summarizing some recent developments, we also seek to bring closer this interesting biomedical application of mathematics to an interdisciplinary readership.

Bhargava, H. N., V. M. Villar, J. Cortijo and E. J. Morcillo. Binding of [3H][D-Ala2, MePhe4, Gly-ol5]enkephalin, [3H][D-Pen2, D-Pen5]enkephalin, and [3H]U-69,593 to airway and pulmonary tissues of normal and sensitized rats. Peptides 18(10) 1603–1608, 1997.—The role of endogenous opioid peptides in the regulation of bronchomotor tone, as well as in the pathophysiology of asthma is uncertain. We have studied the binding of highly selective [3H]labeled ligands of μ-([D-Ala2, MePhe4, Gly-ol5]enkephalin; DAMGO), δ ([D-Pen2, D-Pen5]enkephalin; DPDPE), and κ-(U-69,593) opioid receptors to membranes of trachea, main bronchus, lung parenchyma and pulmonary artery obtained from normal (unsensitized) and actively IgE-sensitized rats acutely challenged with the specific antigen. [3H]DAMGO, [3H]DPDPE and [3H]U-69,593 bound to membranes of normal and sensitized tissues at a saturable, single high-affinity site. The rank order of receptor densities in normal tissues was δ- ≥ κ- ≥ μ-, with lung parenchyma exhibiting the greatest binding capacity for δ- and μ- receptors compared to the other regions examined. The Kd values showed small differences between ligands and regions tested. The μ- and δ-opioid receptor densities were decreased in sensitized main bronchus and lung parenchyma, respectively, compared to normal tissues. By contrast, κ-opioid receptor density was augmented in sensitized lung parenchyma but an increase in Kd values was also observed. These differential changes in the density and affinity of opioid receptor types may be related to alterations in endogenous opioid peptides during the process of sensitization.

Este artículo se encuentra disponible en la siguiente URL: https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0155353 Aims. The main goal of this study was to evaluate the presence of oxidative damage and to quantify its level in gestational diabetes. Methods. Thirty-six healthy women and thirty-six women with gestational diabetes were studied in the three trimesters of pregnancy regarding their levels of oxidative stress markers. These women were diagnosed with diabetes in the second trimester of pregnancy. Blood glucose levels after 100g glucose tolerance test were higher than 190, 165 or 145 mg/dl, 1, 2 or 3 hours after glucose intake. Results. The group of women with gestational diabetes had higher serum malondialdehyde levels, with significant differences between groups in the first and second trimester. The mean values of serum glutathione peroxidase activity in the diabetic women were significantly lower in the first trimester. In the group of women with gestational diabetes there was a negative linear correlation between serum malondialdehyde concentration and glutathione peroxidase activity in the second and third trimester. Conclusions. In this observational and longitudinal study in pregnant women, the alterations attributable to oxidative stress were present before the biochemical detection of the HbA1c increase. Usual recommendations once GD is detected (adequate metabolic control, as well as any other normally proposed to these patients) lowered the concentration of malondialdehyde at the end of pregnancy to the same levels of the healthy controls. Serum glutathione peroxidase activity in women with gestational diabetes increased during the gestational period.

Tetranychus urticae Koch is an important pest affecting citrus, for which biological control has not yet been achieved; therefore, acaricides are commonly used instead. The goal of the work reported in this paper was to measure the efficacy of different new compounds —uracil derivatives— on this mite and conduct a quantitative structure-activity relationship (QSAR) study based on the results obtained, in order to set up a model capable of predicting the acaricidal activity of further new compounds. Some of the tested new products proved highly effective against T. urticae. Besides, topological indices were used as structural descriptors. The result was a topological model consisting of two discriminant functions for distinguishing between active and inactive compounds, and a predictive equation for the adult mortality percentage on the sixth day. This model was then sequentially applied to a large database of compounds with unknown activity against the Tetranychus urticae plague. Finally, a preliminary toxicity study of the most effective novel compounds supports their non-toxicity, performing even better than commercial referents.

The effects of naltrexone injected intravenously (i.v.) on the pharmacological actions and distribution of i.v. injected morphine in brain regions and spinal cord of male Sprague-Dawley rats were determined. Naltrexone (0.625- and 2.5-mg/kg doses) antagonized the analgesic and hyperthermic effects of morphine (10-mg/kg dose). For distribution studies, naltrexone (0.625- and 2.5-mg/kg doses) was co-administered with morphine via indwelling catheters. Rats were sacrificed at various times after drug injection and the concentration of morphine in brain regions (hypothalamus, hippocampus, cortex, pons and medulla, amygdala, midbrain and corpus striatum), spinal cord and serum was determined by radioimmunoassay. The concentration of morphine in various brain regions was found to be time dependent. Initially, at 5 min, the highest concentration of morphine was found in the hypothalamus and the lowest in the striatum. In cortex and spinal cord, the concentration of morphine was significantly higher in comparison to the other brain regions at 30- and 60-min time points. Co-administration of lower dose of naltrexone (0.625 mg/kg) did not significantly alter the distribution of morphine in brain regions and spinal cord with some exceptions. The higher dose of naltrexone (2.5 mg/kg) increased the concentration of morphine in several brain regions and spinal cord. The ratio of the concentration of morphine in brain region or spinal cord to serum was decreased by naltrexone. It is concluded that naltrexone also alters the distribution of morphine in the central nervous system.

Previously it was demonstrated that intravenously administered morphine produced greater analgesic but lower hyperthermic responses to morphine in 24-week-old rats in comparison to 8-week-old rats. The differential pharmacological responses to morphine could not solely be attributed to the pharmacokinetic parameters, namely area under the serum morphine concentration-time curve, serum levels of morphine extrapolated to zero time, half-life, mean residence time, apparent volume of distribution at the steady state, terminal rate constant and total body clearance of morphine in serum. In order to determine whether the differences in pharmacological responses to morphine in rats from two age groups are related to differential distribution of morphine in the central nervous system, in the present study, the time course of the distribution of morphine in brain regions (hypothalamus, hippocampus, cortex, pons and medulla, amygdala, midbrain and corpus striatum), spinal cord and serum following intravenous injection of 10 mg/kg dose to 8- and 24-week-old male Sprague-Dawley rats was determined. Morphine injected intravenously produced a greater analgesic but less intense hyperthermic effect in 24-week-old rats in comparison to 8-week-old rats. In most of the brain regions and spinal cord, with few exceptions, the concentration of morphine was found to be greater in 24-week-old rats than in 8-week-old rats. Similarly, the ratio of the concentration of morphine in brain region or spinal cord to serum was significantly higher in rats from the older age group. The studies demonstrate that the altered pharmacological responses to intravenously administered morphine to rats of differing ages may be related to the higher concentration of morphine in the central nervous system of older rats, which in turn may be related to the differences in the blood-brain barrier to morphine in the two age groups.

PS4α is a high molecular weight peptidoglycan extracted from Mycobacterium vaccae, which has demonstrated considerable antineoplastic activity in-vivo without apparent toxicity. Available for testing in only small quantities, a sensitive in-vivo method for measuring pulse and breathing rates in cannulated rats was applied to this compound at doses of 5, 50 and 500 μg kg−1. Various parameters (mean arterial pressure, maximum transpulmonary pressure, compliance, heart rate, minute volume, respiratory rate and tidal volume) were followed for up to 1 h and demonstrated no significant deviation in the baseline values obtained before injection. This compound at doses up to 500 μg kg−1 had no apparent acute toxicity in rats, but chronic effects at this and higher doses have to be determined by more conventional toxicological methods before proceeding to evaluate PS4α as an antineoplastic agent.

In order to determine the possible contribution of altered distribution of morphine in the morphine tolerance process, the distribution of morphine was studied in brain regions and spinal cord, following its intravenous administration. Male Sprague-Dawley rats were made tolerant to morphine by implanting 6 morphine pellets, each containing 75 mg of morphine base, for 7 days. Seventy-two hours after the removal of the pellets, a time when serum morphine levels were negligible or absent and yet tolerance to the pharmacological effects of morphine was present, morphine (10 mg/kg, i.v.) was injected in placebo and morphine pellet implanted rats. At various times (5, 30, 60, 120 and 360 min) after the injection of morphine, brain regions (hypothalamus, cortex, hippocampus, midbrain, pons and medulla, striatum and amygdala), spinal cord and serum were collected. The level of morphine in the tissues was determined by using a highly sensitive and specific radioimmunoassay (RIA) method. Five minutes after morphine injection, the concentration of morphine was the highest in the hypothalamus and the lowest in amygdala. The concentration of morphine in hypothalamus, pons and medulla, hippocampus and midbrain of morphine tolerant rats was smaller than in placebo pellet implanted rats. The tissue to serum ratio of morphine in the hypothalamus, hippocampus, striatum, midbrain and cortex were also smaller in morphine tolerant than in non-tolerant rats. The concentration of morphine in brain regions with time did not exhibit linearity. At other time intervals like 30 and 60 min, the concentration of morphine in several brain regions and spinal cord was significantly higher in morphine tolerant than in non-tolerant rats.(ABSTRACT TRUNCATED AT 250 WORDS)

The relationship between asthma and opioids has barely been investigated. This study examines whether active sensitization of rats changes the analgesic and thermic effects of intracerebroventricular morphine or the pharmacokinetics of the drug. Morphine (5, 10 and 20 μg) was given intracerebroventricularly to sensitized (active immunization to ovalbumin and Al(OH)3 then airway challenge with ovalbumin after 12 days) and normal (i.e. non-sensitized) male Sprague-Dawley rats. The tail-flick latencies and changes in colon temperature were determined before morphine injection and at 30 min intervals for a period of 300 min afterwards. Results were expressed as the area under the time-response curve. The analgesic and hyperthermic response to morphine for sensitized rats was less than that obtained for normal rats. Cerebrospinal fluid and blood samples were collected periodically for a period of 240 min and morphine levels were determined by a highly sensitive radioimmunoassay. The pharmacokinetic parameters half-life, terminal elimination rate constant and the mean residence time were determined in both cerebrospinal fluid and plasma by non-compartmental analysis. The area under the cerebrospinal fluid concentration-time curve from time zero to infinity was higher for sensitized rats than for normal rats for all three doses of morphine but these differences did not correspond with similar changes in pharmacological responses. In conclusion, the attenuated analgesic and thermic responses to intracerebroventricular morphine in the sensitized rats might be a result of pharmacodynamic alterations rather than to pharmacokinetic changes.

Effects of naltrexone administered intravenously on the pharmacological actions and kinetics of morphine in serum following intravenous administration of morphine were determined in male Sprague-Dawley rats. A 10 mg/kg dose of morphine produced an analgesic response as measured by the tail flick test. Morphine also produced a hyperthermic effect. Naltrexone dose (0.625-2.5 mg/kg)-dependently antagonized the analgesic and hyperthermic effects of morphine. The effect of naltrexone (0.625 and 2.5 mg/kg) on the pharmacokinetic parameters area under the serum morphine concentration time curve (AUC0--infinity), serum levels of morphine extrapolated to zero time (Cmax), half-life (t1/2), mean residence time (MRT), total body clearance (Clt), and volume of distribution at steady state (Vss) of morphine in serum was determined. Naltrexone (0.625 mg/kg) significantly increased AUC0--infinity, Cmax, t1/2 MRT, but decreased the Vss, elimination rate constant (k) and Clt. The higher dose of naltrexone (2.5 mg/kg) produced an increase in the Cmax value of morphine in the serum, but the other pharmacokinetic parameters were unaffected. Since increased morphine concentrations in serum produced by naltrexone cannot explain its antagonistic effects on analgesia and hyperthermia, it is concluded that naltrexone produces its effects by blocking opiate receptors at the appropriate sites. The increases in serum morphine levels by naltrexone may be related to displacement of morphine from protein binding sites and inhibition of morphine metabolism by glucuronyl transferase. This study for the first time demonstrates that in the rat, when morphine and naltrexone are given concurrently, although serum levels of morphine increase, pharmacological effects of morphine are antagonized.(ABSTRACT TRUNCATED AT 250 WORDS)

1. 1. The possible role of pharmacokinetics of morphine in the development of tolerance to the analgesic and hyperthermic effects of morphine was studied in the rat. 2. 2. Male Sprague-Dawley rats were made tolerant to morphine by implanting 6 morphine pellets each containing 75 mg of morphine base for 7 days. The assessment of the degree of tolerance to morphine and pharmacokinetic parameters were done 72 hr after pellet removal. 3. 3. Tolerance developed to both the analgesic and hyperthemic effects of morphine as evidenced by decreased responses to morphine in morphine pellet implanted rats compared with placebo pellet implanted rats. 4. 4. The pharmacokinetic parametrs, AUC0→∞, Cmax, t 1 2 , k, MRT, Vss and Clt were determined after injecting 5 and 10 mg/kg doses of morphine intravenously to placebo and morphine pellet implanted rats and using a highly sensitive and specific RIA method to quantitate serum levels of morphine. For a 5 mg/kg dose of morphine, the AUC0→∞ and t 1 2 in morphine pellet implanted rats were significantly higher than in placebo pellet implanted rats, but the k value was lower. The other pharmacokinetic parameters for morphine in the two treatment groups did not differ. For 10 mg/kg dose, the only change was an increase in the MRT in morphine tolerant rats when compared to nontolerant rats. 5. 5. The results establish that the development of tolerance to the analgesic and hyperthermic effects of morphine is not related to pharmacokinetics of morphine in serum but may be related to modification of receptor systems in the central nervous system.

1. In order to determine whether the degree of tolerance to and physical dependence on morphine induced by pellet implantation procedure in the rat depends on the dose used and the kinetic parameters, the effect of implantation of different number of pellets on tolerance-dependence and elimination kinetics of morphine from serum was determined. 2. Male Sprague-Dawley rats were implanted subcutaneously with pellets. Each pellet contained 75 mg of morphine free base. Three schedules of implantation were used. They included 2 pellets during a 3-day period ( 2 3 ), 4 pellets during a 3-day period ( 4 3 ) and 6 pellets during a 7-day period ( 6 7 ). Placebo pellets which did not contain morphine were implanted in rats which served as controls. 3. The degree of tolerance to and physical dependence on morphine increased as the number of morphine pellets implanted increased. 4. In separate groups of rats implanted with pellets, elimination kinetics of morphine was studied using radioimmunoassay. The kinetic parameters were: area under serum morphine concentration time curve (AUC0→∞), serum concentration of morphine extrapolated to time zero (Cmax), half-life ( t 1 2 ), elimination rate constant (k), mean residence time (MRT) and total body clearance (Clt). 5. The AUC0→∞ and Cmax increased in proportion to the number of pellets implanted. The t 1 2 , k, MRT and Clt values for 2 3 and 4 3 schedules did not differ, but for 6 7 schedule were significantly different from the other two schedules. The degree of tolerance to and physical dependence on morphine was directly related to the AUC0→∞ and Cmax. The longer t 1 2 and MRT and lower Clt and k values in 6 7 schedule may reflect a saturation of glucuronic acid transferase, the main enzyme metabolizing morphine in the liver, and may account for the greater degree of tolerance and physical dependence.

The analgesic and thermic responses to morphine (5 and 10 mg/kg) injected intravenously to 8- and 24-week-old male Sprague-Dawley rats were determined. Greater analgesic and lower hyperthermic responses to morphine in 24-week-old rats in comparison to 8-week-old rats were observed. The pharmacokinetic parameters of morphine administered intravenously were also determined. Cmax for 5 and 10 mg/kg doses of morphine were smaller in 24-week-old rats in comparison to 8-week-old rats; however, AUC0----infinity was smaller only for 5 mg/kg dose. For 10 mg/kg dose, mean residence time (MRT) and the apparent steady state volume of distribution (Vss) for the older rats were higher than for the younger ones, but for 5 mg/kg dose the values did not differ. The enhanced responses to morphine in older age group of rats for 5 mg/kg dose cannot be explained solely on the basis of pharmacokinetics. However, for 10 mg/kg dose of morphine, the greater responses in 24-week-old rats could probably be related to increases in MRT and Vss. Factors other than serum kinetics, like kinetics of morphine in the brain as well as the brain opiate receptors, may also be involved in the differential effects of morphine in rats of different ages.