Exposure to varenicline protects against locomotor alteration in a MPTP mouse model of Parkinson's disease.

The beneficial effects of drugs that act via nicotinic acetylcholine receptors (nAChRs) on Parkinson's disease (PD) symptomatology may explain the negative correlation between cigarette smoking and risk of this neurological condition. Varenicline, an α4ß2 nAChR partial agonist approved for smoking cessation treatments, could be valuable for PD treatment. Here, we investigated varenicline effects in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) PD mouse model. From postnatal day (PN) 90 to PN119, male C57BL/6 mice were exposed daily to varenicline (2 mg/kg) by gavage. After that, MPTP was injected (30 mg/kg, ip) once a day for five days. At PN125, locomotor and anxiety-like effects were assessed with the open field test. At PN126, immobile behavior was assessed with the forced swimming test. At PN127, the frontal cerebral cortex was collected to evaluate dopamine and DOPAC levels. To verify whether varenicline was protective during the MPTP insult, a separate group of MPTP animals received varenicline from PN90 to PN124. MPTP reduced cortical dopamine content and increased dopamine turnover. Those effects were not reversed by varenicline treatment. Interestingly, varenicline reversed the MPTP-induced hyperactivity in the open field. Both maintenance of varenicline treatment during MPTP exposure or its interruption before MPTP exposure elicited similar results. No alterations were observed in anxiety-like behavior or in immobility time. Altogether, these findings suggested that varenicline treatment reduced the MPTP-induced hyperactivity, but did not protect against dopaminergic damage. Based on this partial protective effect, varenicline could exert neuroprotective effects on circuits that control motor activity in PD.

Exposure to varenicline protects against locomotor alteration in a MPTP mouse model of Parkinson's disease

The beneficial effects of drugs that act via nicotinic acetylcholine receptors (nAChRs) on Parkinson's disease (PD) symptomatology may explain the negative correlation between cigarette smoking and risk of this neurological condition. Varenicline, an α4β2 nAChR partial agonist approved for smoking cessation treatments, could be valuable for PD treatment. Here, we investigated varenicline effects in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) PD mouse model. From postnatal day (PN) 90 to PN119, male C57BL/6 mice were exposed daily to varenicline (2 mg/kg) by gavage. After that, MPTP was injected (30 mg/kg, ip) once a day for five days. At PN125, locomotor and anxiety-like effects were assessed with the open field test. At PN126, immobile behavior was assessed with the forced swimming test. At PN127, the frontal cerebral cortex was collected to evaluate dopamine and DOPAC levels. To verify whether varenicline was protective during the MPTP insult, a separate group of MPTP animals received varenicline from PN90 to PN124. MPTP reduced cortical dopamine content and increased dopamine turnover. Those effects were not reversed by varenicline treatment. Interestingly, varenicline reversed the MPTP-induced hyperactivity in the open field. Both maintenance of varenicline treatment during MPTP exposure or its interruption before MPTP exposure elicited similar results. No alterations were observed in anxiety-like behavior or in immobility time. Altogether, these findings suggested that varenicline treatment reduced the MPTP-induced hyperactivity, but did not protect against dopaminergic damage. Based on this partial protective effect, varenicline could exert neuroprotective effects on circuits that control motor activity in PD.

Concurrent maternal and pup postnatal tobacco smoke exposure in Wistar rats changes food preference and dopaminergic reward system parameters in the adult male offspring.

Children from pregnant smokers are more susceptible to become obese adults and to become drug or food addicts. Drugs and food activate the mesolimbic reward pathway, causing a sense of pleasure that induces further consumption. Here, we studied the relationship between tobacco smoke exposure during lactation with feeding, behavior and brain dopaminergic reward system parameters at adulthood. Nursing Wistar rats and their pups were divided into two groups: tobacco smoke-exposed (S: 4times/day, from the 3rd to the 21th day of lactation), and ambient air-exposed (C). On PN175, both offspring groups were subdivided for a food challenge: S and C that received standard chow (SC) or that chose between high-fat (HFD) and high-sucrose diets (HSDs). Food intake was recorded after 30min and 12h. Offspring were tested in the elevated plus maze and open field on PN178-179; they were euthanized for dopaminergic analysis on PN180. SSD (self-selected diet) animals presented a higher food intake compared to SC ones. S-SSD animals ate more than C-SSD ones at 30min and 12h. Both groups preferred the HFD. However, S-SSD animals consumed relatively more HFD than C-SSD at 30min. No behavioral differences were observed between groups. S animals presented lower tyrosine hydroxylase (TH) content in the ventral tegmental area, lower TH, dopaminergic receptor 2, higher dopaminergic receptor 1 contents in the nucleus accumbens and lower OBRb in hypothalamic arcuate nucleus. Tobacco-smoke exposure during lactation increases preference for fat in the adult progeny possibly due to alterations in the dopaminergic system.

Endocrine effects of tobacco smoke exposure during lactation in weaned and adult male offspring.

Children from pregnant smokers show more susceptibility to develop obesity in adult life. Previously, we failed to demonstrate a program for obesity in rat offspring only when the mothers were exposed to tobacco smoke during lactation. Here, we studied the short- and long-term effects of smoke exposure (SE) to both dams and their pups during lactation on endocrine and metabolic parameters. For this, we designed an experimental model where nursing rats and their pups were divided into two groups: SE group, exposed to smoke in a cigarette smoking machine (four times/day, from the third to the 21st day of lactation), and group, exposed to filtered air. Pups were killed at 21 and 180 days. At weaning, SE pups showed lower body weight (7%), length (5%), retroperitoneal fat mass (59%), visceral adipocyte area (60%), and higher subcutaneous adipocyte area (95%) with hypoinsulinemia (-29%), hyperthyroxinemia (59%), hypercorticosteronemia (60%), and higher adrenal catecholamine content (+58%). In adulthood, SE offspring showed higher food intake (+10%), body total fat mass (+50%), visceral fat mass (retroperitoneal: 55%; mesenteric: 67%; and epididymal: 55%), and lower subcutaneous adipocyte area (24%) with higher serum glucose (11%), leptin (85%), adiponectin (1.4-fold increase), total triiodothyronine (71%), free thyroxine (57%), TSH (36%), triglycerides (65%), VLDL cholesterol (+66%), and HDL cholesterol (91%) levels and lower corticosteronemia (41%) and adrenal catecholamine content (57%). Our present findings suggest that tobacco SE to both dams and their pups during lactation causes malnutrition in early life that programs for obesity and hormonal and metabolic disturbances in adulthood, only if the pups are submitted to the same smoke environment as the mother.

Maternal tobacco smoke exposure during lactation inhibits catecholamine production by adrenal medullae in adult rat offspring.

Previously, we have shown that maternal smoke exposure during lactation, even when pups are not exposed, affects biochemical profiles in the offspring at weaning, eliciting lower body adiposity, hyperinsulinemia, hypocorticosteronemia and lower adrenal catecholamine content. However, the future impact of tobacco exposure is still unknown. As postnatal nicotine exposure causes short- and long-term effects on pups' biochemistry and endocrine profiles, we have now evaluated some endocrine and metabolic parameters of the adult offspring whose mothers were tobacco exposed during lactation. For this, from day 3 to 21 of lactation, rat dams were divided in: 1) SE group, cigarette smoke-exposed (1.7 mg nicotine/cigarettes for 1 h, 4 times/day, daily), without their pups, and 2) C group, exposed to air, in the same conditions. Offspring were killed at 180-days-old. Body weight and food intake were evaluated. Blood, white adipose tissue, adrenal, and liver were collected. All significant data were p<0.05. The adult SE offspring showed no change in body weight, cumulative food intake, serum hormone profile, serum lipid profile, or triglycerides content in liver. However, in adrenal gland, adult SE offspring showed lower catecholamine content ( - 50%) and lower tyrosine hydroxylase protein expression ( - 56%). Despite the hormonal alterations during lactation, tobacco smoke exposure through breast milk only programmed the adrenal medullary function at adulthood and this dysfunction can have consequence on stress response. Thus, an environment free of smoke during lactation period is essential to improve health outcomes in adult offspring.

Effects of tobacco smoke exposure during lactation on nutritional and hormonal profiles in mothers and offspring.

Exposure to tobacco smoke is related to changes in energy balance regulation and several endocrine dysfunctions. Previously, we showed that maternal nicotine (the main addictive compound of tobacco) exposure exclusively during lactation affects biochemical profiles in mothers, milk, and pups. As the possible consequences for mothers and offspring of maternal smoking during lactation are still unknown, we evaluated the effects of tobacco smoke exposure on nutritional, biochemical, and hormonal parameters in dams and pups at weaning. After 72 h from birth, lactating rats were divided into two groups: smoke-exposed (S) in a cigarette-smoking machine, 4 × 1 h per day throughout the lactation period without pups; control (C), rats were treated the same as the experimental group but exposed to filtered air. Dams and pups were killed at weaning (21 days of lactation). Body weight and food intake were evaluated. Milk, blood, visceral fat, adrenal, and carcass were collected. S dams showed hyperprolactinemia (+50%), hypoinsulinemia (-40%), hypoleptinemia (-46%), as well as lower triglycerides (-53%) and very low-density lipoprotein cholesterol (-50%). Milk of S dams had higher lactose (+52%) and triglycerides (+78%). S pups presented higher body protein (+17%), lower total (-24%) and subcutaneous fat contents (-25%), hypoglycemia (-11%), hyperinsulinemia (+28%), hypocorticosteronemia (-40%), lower adrenal catecholamine content (-40%), hypertriglyceridemia (+34%), higher high-density lipoprotein cholesterol (+16%), and lower low-density lipoprotein cholesterol (-45%). In conclusion, tobacco smoke exposure leads to changes in nutritional, biochemical, and hormonal parameters in dams and, passively through the milk, may promote several important metabolic disorders in the progeny.

Neonatal nicotine exposure alters leptin signaling in the hypothalamus-pituitary-thyroid axis in the late postnatal period and adulthood in rats.

AIMS: Postnatal nicotine exposure causes precocious primary hypothyroidism and programs for overweight, hyperleptinemia and secondary hypothyroidism in adulthood. As leptin and thyroid hormones share the ability to increase energy expenditure, we studied the effects of maternal nicotine exposure during lactation on the leptin signaling in the hypothalamus-pituitary-thyroid axis of suckling and adult offspring. MAIN METHODS: Two days after delivery, osmotic minipumps were implanted in lactating rats, and nicotine (NIC, 6 mg/kg/day s.c.) or saline (C) was administered for 14days. Offspring were killed at 15 and 180 days-old. Proteins belonging to leptin signaling were analyzed by Western blot. Significant differences had p<0.05. KEY FINDINGS: In the hypothalamus, NIC offspring showed higher OB-R and pSTAT-3 content (+58%,+1.34x) at 15 days, and lower OB-R, JAK-2 and pSTAT-3 (-61%, -42%, -56%) at 180 days. In the pituitary gland, NIC offspring showed lower JAK-2 content (-52%) at 15 days, but no differences in adulthood. In the thyroid gland, the NIC group presented lower OB-R, JAK-2 and STAT-3 (-44%, -50%, -47%) and higher pSTAT-3 expression (+80%) at 15 days. At 180 days-old, NIC offspring presented higher thyroid OB-R (+1.54x) and lower pSTAT-3 content (-34%). SIGNIFICANCE: Neonatal primary hypothyroidism induced by maternal nicotine exposure during lactation may be partially explained by decreased leptin signaling in the thyroid, though the early stimulation of the central leptin pathway did not prevent the thyroid dysfunction. Long-term effects of postnatal nicotine exposure on leptin signaling in the hypothalamus and thyroid appear to involve central and peripheral leptin resistance in adulthood.

Nicotine exposure affects mother's and pup's nutritional, biochemical, and hormonal profiles during lactation in rats.

We have shown that maternal nicotine exposure during lactation has long-lasting effects on body adiposity and hormonal status of rat offspring. Here, we studied the nutritional and hormonal profiles in this experimental model. Two days after birth, osmotic minipumps were implanted in lactating rats divided into two groups: NIC - continuous s.c. infusions of nicotine (6 mg/kg per day) for 14 days and C - saline. Dams and pups were killed at 15 and 21 days of lactation. Body weight and food intake were evaluated. Milk, blood, visceral fat, carcass, and adrenal gland were collected. All the significant data were P<0.05. At the end of nicotine exposure (15 days), dams presented higher milk production, hyperprolactinemia, and higher serum high-density lipoprotein cholesterol (HDL-C). Milk from NIC dams had higher lactose concentration and energy content. After nicotine withdrawal (21 days), dams showed lower food intake and hyperleptinemia. The 15-day-old NIC pups presented higher total body fat, higher HDL-C, serum leptin, serum corticosterone, and adrenal catecholamine content, but lower tyrosine hydroxylase protein levels. The 21-day-old NIC pups had higher body protein content and serum globulin. Thus, maternal nicotine exposure during lactation results in important changes in nutritional, biochemical, and hormonal parameters in dams and offspring. The pattern of these effects is clearly distinct when comparing the nicotine-exposed group to the withdrawal group, which could be important for the programming effects observed previously.

Hippocampal increased cell death and decreased cell density elicited by nicotine and/or ethanol during adolescence are reversed during drug withdrawal.

We have recently identified hippocampal cell death and reduced neuronal and glial cells densities during adolescent nicotine and ethanol exposures and outcomes reduced in severity when nicotine and ethanol are co-administered during this developmental period. In the present study, we investigated the effects of adolescent nicotine and/or ethanol withdrawal on the following regions of the hippocampus: Granular layer of the Dentate Gyrus (GrDG), Molecular layer (Mol), CA1, CA2 and CA3. From the 30th to the 45th postnatal day (PN30-PN45), C57BL/6 male and female mice were exposed to nicotine free base (NIC) and/or ethanol (ETOH). Four groups were analyzed: (1) concomitant NIC (50 microg/ml in 2% saccharin to drink) and ETOH (25%, 2 g/kg i.p. injected every other day) exposure; (2) NIC exposure; (3) ETOH exposure; (4) vehicle. We evaluated cell degeneration (TUNEL assay), neuronal and glial densities (optical Disector) and region thicknesses two (PN47) and five (PN50) days post-exposure. On PN47, there were increases in the number of TUNEL+ cells in most hippocampal regions of both ETOH and NIC groups. In the NIC+ETOH group there were less severe effects. These results were paralleled by reductions in neuronal and glial cells densities for all treatment groups. In contrast, on PN50, ethanol and/or nicotine withdrawal were associated with compensatory reductions in TUNEL+ cells in all hippocampal regions. These results were paralleled by a reversal of effects on neuronal and glial densities so that there were no longer differences between groups. There were no effects on region thicknesses. These results suggest that deleterious effects of nicotine and/or ethanol are reversed during prolonged withdrawal.

Short- and long-term effects of maternal nicotine exposure during lactation on body adiposity, lipid profile, and thyroid function of rat offspring.

Epidemiological studies show a higher prevalence of obesity in children from smoking mothers and smoking may affect human thyroid function. To evaluate the mechanism of smoking as an imprinting factor for these dysfunctions, we evaluated the programming effects of maternal nicotine (NIC) exposure during lactation. Two days after birth, osmotic minipumps were implanted in lactating rats, divided into: NIC (6 mg/kg per day s.c.) for 14 days; Control - saline. All the significant data were P<0.05 or less. Body weight was increased from 165 days old onwards in NIC offspring. Both during exposure (at 15 days old) and in adulthood (180 days old), NIC group showed higher total fat (27 and 33%). In addition, NIC offspring presented increased visceral fat and total body protein. Lipid profile was not changed in adulthood. Leptinemia was higher at 15 and 180 days old (36 and 113%), with no changes in food intake. Concerning the thyroid status, the 15-days-old NIC offspring showed lower serum-free tri-iodothyronine (FT(3)) and thyroxine (FT(4)) with higher TSH. The 180-days-old NIC offspring exhibited lower TSH, FT(3), and FT(4)). In both periods, liver type 1 deiodinase was lower (26 and 55%). We evidenced that NIC imprints a neonatal thyroid dysfunction and programs for a higher adiposity, hyperleptinemia, and secondary hypothyroidism in adulthood. Our study identifies lactation as a critical period to NIC programming for obesity, with hypothyroidism being a possible contributing factor.

Combined exposure to nicotine and ethanol in adolescent mice: effects on the central cholinergic systems during short and long term withdrawal.

Relapse to drug use is a major public health problem. In this sense, understanding the biological substrates that are affected during withdrawal may provide information to prevent relapse. Both smoking and alcoholic beverage consumption usually begin during adolescence, however, little is known about the basic neurobiology of the combined adolescent exposure, particularly during withdrawal. Since nicotine is a cholinergic agonist and it has been shown that ethanol interferes with nicotinic acetylcholine receptors (nAChRs), the current study focused on the effects of drug withdrawal on the central cholinergic system. From the 30th to the 45th postnatal day (PN), C57BL/6 male and female mice were exposed to nicotine free base (NIC) and/or ethanol (ETOH). Four groups were analyzed: (1) concomitant NIC (50 microg/ml in 2% saccharin to drink) and ETOH (25%, 2 g/kg i.p. injected every other day) exposure; (2) NIC exposure; (3) ETOH exposure; (4) vehicle. We assessed nAChR binding, choline acetyltransferase (ChAT) activity and [(3)H]hemicholinium-3 (HC-3) binding in the cerebral cortex and midbrain of mice at short (PN50) and long term (PN75) withdrawal. NIC and NIC+ETOH promoted nAChR upregulation during a short-term withdrawal. NIC short-term withdrawal elicited an increase in ChAT activity that was reversed by ETOH withdrawal. In addition, NIC+ETOH elicited a decrease in ChAT activity at long term withdrawal. Regarding HC-3, ETOH and NIC+ETOH promoted a decrease that persisted at long-term withdrawal. The present study provides experimental evidence that nicotine and ethanol during adolescence interact resulting in cholinergic system alterations during withdrawal.

The effect of corpus callosum agenesis on neocortical thickness and neuronal density of BALB/cCF mice.

We used acallosal and normal adult BALB/cCF mice to test the hypothesis that the development of the corpus callosum is relevant for the establishment of a normal structure of the neocortex. Neuronal density and thickness of individual layers were analyzed in neocortical regions with abundant callosal connections (area 6 and the 17/18a border) and in the relatively acallosal area 17. In area 6, acallosal mice exhibited a total neocortical thickness smaller than that of normal mice, as well as thinner layers II+III and IV. Similar data were obtained at the 17/18a border, where the total thickness of the cortex and of layers II+III was smaller in the acallosal mice than in normal ones. In contrast, no significant thickness differences were documented in area 17 of acallosal versus normal mice. The quantitative data obtained in the analyzed neocortical regions did not show differences in neuronal density between acallosal and normal mice. The reduced cortical thickness, associated with the comparatively normal neuronal density in neocortical regions which normally have abundant callosal connections, provides indirect indication of a reduction in the number of cortical neurons in acallosal mice. This assumption was also supported by the lack of evidence of neocortical alterations in the acallosal area 17. The present findings suggest that during development neocortical neurons destined to receive a massive callosal input may die as a result of lack of afferents. Altogether the present data indicate that the input provided by callosal axons is necessary for a normal development of the neocortex.

Time course of the effects of prenatal gamma irradiation on the dorsal lateral geniculate nucleus of Swiss mice.

A previous study reported that adult mice irradiated at the 16th embryonic day present a severe neuronal number reduction in the dorsal lateral geniculate thalamic nucleus. In the present study, we investigated the time course of the effects of prenatal irradiation on this thalamic nucleus. One day after irradiation, a great number of pyknotic figures were seen mainly in the cerebral proliferative zones. In the geniculate nucleus, only scattered pyknotic figures were identified. On the first week after birth, the geniculate nucleus presented frequent pyknotic figures. From five days after birth onwards, a severe shrinkage of the occipital cortex and a great reduction in the geniculate nucleus neuronal number were found. On the second week after birth this neuronal number reduction reached as high as 75%. At each postnatal analyzed age, severe volumetric geniculate nucleus shrinkage was combined to non-significant neuronal density variations. The presence of few pyknotic figures in the geniculate nucleus one day after irradiation and its delayed neuronal loss indicate an indirect effect of irradiation. We suggest that the effect upon the geniculate nucleus is secondary to the damage of the occipital cortex. A possible interpretation for thalamic neuronal loss is that geniculate neurons fail to establish cortical arbors after major target loss. In this case, the loss of trophic support should also be considered.

Influences of handedness and gender on the grooved pegboard test.

We studied performance on the Grooved Pegboard Test upon repeated trials and transfer of training between the hands in the first trial. The classification of handedness was based on the writing hand. We employed three trials for each hand and two different protocols for the order in which the hands started the test. For the three trials combined, women were faster than men. From the first to the second trial, there was an improvement in performance for both sexes. Within the first trial, sex differences reached significance and the protocol interacted with handedness. In this trial, only left-handed men were found to benefit from previous opposite-hand performance. It is speculated that a larger corpus callosum in left-handed men allows for the greater transfer of training between the hands.

Effects of prenatal gamma irradiation on the development of the corpus callosum of Swiss mice.

The temporal sequence of events related to the effects of prenatal gamma irradiation on the development of the corpus callosum and cerebral cortex was studied in Swiss mice. Pregnant females on gestational day 16 were exposed to a 60Co source receiving total doses of 2 or 3 Gy. The offspring were analyzed at both prenatal and postnatal days. One day after irradiation, a great number of pyknotic figures was seen along the whole extension of the cerebral wall, especially in the proliferative zones. At perinatal ages, the thickness of the proliferative zones was reduced and the glial sling was never identified. From 5 days after birth onwards, we observed a severe shrinkage of layers II + III and IV. The majority of the irradiated mice were totally acallosal (particularly when the 3 Gy dose was used), but some animals presented callosal remnants. These remnants were identified above the ventral hippocampal commissure, except for two animals in which a larger callosal remnant extended from the columns of the fornix to the dorsal hippocampal commissure. The presence of callosal remnants in animals irradiated with 3 Gy was dependent on the age at which the animals were analyzed since remnants were observed in some animals analyzed at perinatal ages, but never in older animals. Callosal defects can be explained at least by three factors: (1) Death of a great part of callosal neurons located at layer III. (2) Postnatal axonal elimination. (3) Absence of the glial sling. The callosal agenesis in the absence of the glial sling indicates that this structure may play a crucial role in guiding callosal axons. However, the presence of callosal remnants indicates that surviving callosal axons can use structures other than the sling to cross the midplane. Our data indicate that axons of the middle portion of the callosum can cross the midplane using the ventral hippocampal commissure as a guide. Additionally, the dorsal hippocampal commissure may play a role in directing axons of the posterior part of the corpus callosum.