Metformin effectively restores the HPA axis function in diet-induced obese rats.

INTRODUCTION: The hypothalamo-pituitary-adrenal (HPA) axis is perturbed in obesity. We previously reported presence of leptin resistance in the brainstem and uncoupling between central noradrenergic tone and the HPA axis in obesity-prone (DIO) rats. Metformin is shown to lower body weight and adiposity, but the underlying mechanism is unclear. We hypothesized that this is associated with restored HPA axis function. METHODS: Adult male DIO rats were placed on either a regular chow or HF diet for 7 weeks. Starting week 4, the animals were given either a low dose (60 mg/kg) or high dose (300 mg/kg) of metformin in drinking water. In addition to body weight and feeding, we examined different arms of the HPA axis to test if metformin can reinstate its function and coupling. To understand potential mechanisms, leptin signaling in the brainstem and circulating free fatty acid levels were also assessed. RESULTS: Metformin treatment lowered weight gain, fat mass, caloric intake, and serum leptin levels. HPA axis activity as determined by corticotropin-releasing hormone in the median eminence and serum corticosterone was decreased by metformin in a dose-dependent manner, and so was norepinephrine (NE) in the paraventricular nucleus. Importantly, metformin completely normalized the NE-HPA axis uncoupling. While brainstem pSTAT-3 and SOCS-3, key markers of leptin signaling, were not different between groups, circulating saturated and unsaturated free fatty acids were reduced in HF-fed, metformin-treated animals. CONCLUSIONS: These findings suggest that oral metformin can successfully correct HPA axis dysfunction that is associated with lowered circulating free fatty acids in DIO rats, thereby uncovering a novel effect of metformin in the treatment of obesity.

Evaluation of the Central Effects of Systemic Lentiviral-Mediated Leptin Delivery in Streptozotocin-Induced Diabetic Rats.

Type 1 diabetes (T1D) is characterized by hyperphagia, hyperglycemia and activation of the hypothalamic-pituitary-adrenal (HPA) axis. We have reported previously that daily leptin injections help to alleviate these symptoms. Therefore, we hypothesized that leptin gene therapy could help to normalize the neuroendocrine dysfunction seen in T1D. Adult male Sprague Dawley rats were injected i.v. with a lentiviral vector containing the leptin gene or green fluorescent protein. Ten days later, they were injected with the vehicle or streptozotocin (STZ). HPA function was assessed by measuring norepinephrine (NE) levels in the paraventricular nucleus (PVN) and serum corticosterone (CS). Treatment with the leptin lentiviral vector (Lepvv) increased leptin and insulin levels in non-diabetic rats, but not in diabetic animals. There was a significant reduction in blood glucose levels in diabetic rats due to Lepvv treatment. Both NE levels in the PVN and serum CS were reduced in diabetic rats treated with Lepvv. Results from this study provide evidence that leptin gene therapy in STZ-induced diabetic rats was able to partially normalize some of the neuroendocrine abnormalities, but studies with higher doses of the Lepvv are needed to develop this into a viable option for treating T1D.

Chronic estradiol exposure - harmful effects on behavior, cardiovascular and reproductive functions.

Estradiol (E2) is a female hormone that is produced largely by the ovaries, but also by the adrenal glands, fat and liver. It is present in the circulation of both males and females. Many studies in the literature have described how E2 is beneficial to the body in terms of preventing bone loss, affording protection in ischemia reperfusion injury, relieving symptoms of menopause, maintaining vaginal health and helping with ovarian failure or hypogonadism. Beneficial effects on the brain have been reported to include protection against memory loss, neuronal degeneration, changes in cognition, mood and behavior. However, the effects of E2 exposure on the neuroendocrine system have not been understood completely. This is because differences in doses, preparation and duration of exposure have produced variable results ranging from beneficial, to no change, or to detrimental. Studies in our lab over the last few years have shown that chronic exposures to low levels of E2 in young rats can produce specific effects on the neuroendocrine system. We have observed that these exposures can induce reproductive senescence, hypertension, anxiety-like behavior and cause degenerative changes in specific neuronal populations leading to hyperprolactinemia. The purpose of the review is to present evidence from the literature for these effects and to discuss the underlying molecular mechanisms.

Differential effects of prenatal stress on metabolic programming in diet-induced obese and dietary-resistant rats.

Stress during pregnancy is a known contributing factor for the development of obesity in the offspring. Since maternal obesity is on the rise, we wanted to identify the effects of prenatal stress in the offspring of diet-induced obese (DIO) rats and compare them with the offspring of dietary-resistant (DR) rats. We hypothesized that prenatal stress would make both DIO and DR offspring susceptible to obesity, but the effect would be more pronounced in DIO rats. Pregnant DIO and DR rats were divided into two groups: nonstressed controls (control) and prenatal stress (subjected to restraint stress, three times/day from days 14 to 21 of gestation). After recording birth weight and weaning weight, male offspring were weaned onto a chow diet for 9 wk and shifted to a high-fat (HF) diet for 1 wk. At the end of the 10th wk the animals were euthanized, and visceral adipose mass, blood glucose, serum insulin, and C-peptide levels were measured. Prenatal stress resulted in hyperinsulinemia and higher C-peptide levels without altering caloric intake, body weight gain, or fat mass in the DIO offspring after 1 wk of HF intake, but not in DR offspring. To determine the mechanism underlying the hyperinsulinemia, we measured the levels of CEACAM1 that are responsible for insulin clearance. CEACAM1 levels in the liver were reduced in prenatally stressed DIO offspring after the HF challenge, suggesting that preexisting genetic predisposition in combination with prenatal stress increases the risk for obesity in adulthood, especially when offspring are fed a HF diet.

Role of cytokines and reactive oxygen species in brain aging.

Aging is a complex process that produces profound effects on the brain. Although a number of external factors can promote the initiation and progression of brain aging, peripheral and central changes in the immune cells with time, also play an important role. Immunosenescence, which is an age-associated decline in immune function and Inflammaging, a low-grade inflammatory state in the aging brain contribute to an elevation in cytokine and reactive oxygen species production. In this review, we focus on the pro-inflammatory state that is established in the brain as a consequence of these two phenomena and the resulting detrimental changes in brain structure, function and repair that lead to a decline in central and neuroendocrine function.

Prenatal exposure to bisphenol A and/or diethylhexyl phthalate alters stress responses in rat offspring in a sex- and dose-dependent manner.

Background: Prenatal exposures to endocrine disrupting chemicals (EDCs) are correlated with adverse behavioral outcomes, but the effects of combinations of these chemicals are unclear. The aim of this study was to determine the dose-dependent effects of prenatal exposure to EDCs on male and female behavior. Methods: Pregnant Sprague-Dawley rats were orally dosed with vehicle, bisphenol A (BPA) (5 µg/kg body weight (BW)/day), low-dose (LD) diethylhexyl phthalate (DEHP) (5 µg/kg BW/day), high-dose (HD) DEHP (7.5 mg/kg BW/day), a combination of BPA and LD-DEHP (B + D (LD)), or a combination of BPA and HD-DEHP (B + D (HD)) on gestational days 6-21. Adult offspring were subjected to the Open Field Test (OFT), Elevated Plus Maze (EPM), and Shock Probe Defensive Burying test (SPDB) in adulthood. Body, adrenal gland, and pituitary gland weights were collected at sacrifice. Corticosterone (CORT) was measured in the serum. Results: Female EDC-exposed offspring showed anxiolytic effects in the OFT, while male offspring were unaffected. DEHP (HD) male offspring demonstrated a feminization of behavior in the EPM. Most EDC-exposed male offspring buried less in the SPDB, while their female counterparts showed reduced shock reactivity, indicating sex-specific maladaptive alterations in defensive behaviors. Additionally, DEHP (LD) males and females and B + D (LD) females displayed increased immobility in this test. DEHP (LD) alone and in combination with BPA led to lower adrenal gland weights, but only in male offspring. Finally, females treated with a mixture of B + D (HD) had elevated CORT levels. Conclusion: Prenatal exposure to BPA, DEHP, or a mixture of the two, affects behavior, CORT levels, and adrenal gland weights in a sex- and dose-dependent manner.

Branched-Chain Amino Acids Are Linked with Alzheimer's Disease-Related Pathology and Cognitive Deficits.

Alzheimer's disease (AD) is an irreversible neurodegenerative disorder with a complex pathophysiology. Type 2 diabetes (T2D) is a strong risk factor for AD that shares similar abnormal features including metabolic dysregulation and brain pathology such as amyloid and/or Tau deposits. Emerging evidence suggests that circulating branched-chain amino acids (BCAAs) are associated with T2D. While excess BCAAs are shown to be harmful to neurons, its connection to AD is poorly understood. Here we show that individuals with AD have elevated circulating BCAAs and their metabolites compared to healthy individuals, and that a BCAA metabolite is correlated with the severity of dementia. APPSwe mouse model of AD also displayed higher plasma BCAAs compared to controls. In pursuit of understanding a potential causality, BCAA supplementation to HT-22 neurons was found to reduce genes critical for neuronal health while increasing phosphorylated Tau. Moreover, restricting BCAAs from diet delayed cognitive decline and lowered AD-related pathology in the cortex and hippocampus in APP/PS1 mice. BCAA restriction for two months was sufficient to correct glycemic control and increased/restored dopamine that were severely reduced in APP/PS1 controls. Treating 5xFAD mice that show early brain pathology with a BCAA-lowering compound recapitulated the beneficial effects of BCAA restriction on brain pathology and neurotransmitters including norepinephrine and serotonin. Collectively, this study reveals a positive association between circulating BCAAs and AD. Our findings suggest that BCAAs impair neuronal functions whereas BCAA-lowering alleviates AD-related pathology and cognitive decline, thus establishing a potential causal link between BCAAs and AD progression.

Chronic estradiol exposure induces oxidative stress in the hypothalamus to decrease hypothalamic dopamine and cause hyperprolactinemia.

Estrogens are known to cause hyperprolactinemia, most probably by acting on the tuberoinfundibular dopaminergic (TIDA) system of the hypothalamus. Dopamine (DA) produced by TIDA neurons directly inhibits prolactin secretion and, therefore, to stimulate prolactin secretion, estrogens inhibit TIDA neurons to decrease DA production. However, the mechanism by which estrogen produces this effect is not clear. In the present study, we used a paradigm involving chronic exposure to low levels of estradiol-17ß (E(2)) to mimic prolonged exposures to environmental and endogenous estrogens. We hypothesized that chronic exposure to low levels of E(2) induces oxidative stress in the arcuate nucleus (AN) of the hypothalamus that contains TIDA neurons and causes nitration of tyrosine hydroxylase (TH), the rate-limiting enzyme in the synthesis of DA. This results in a significant decrease in DA and consequently, hyperprolactinemia. To investigate this, adult, intact female cycling rats were implanted with slow-release E(2) pellets (20 ng/day) for 30, 60, or 90 days and were compared with old (16-18 mo old) constant estrous (OCE) rats. Chronic E(2) exposure significantly increased the expression of glial fibrillary acidic protein and the concentrations of interleukin-1ß (IL-1ß) and nitrate in the AN that contains perikarya of TIDA neurons and increased nitration of TH in the median eminence (ME) that contains the terminals. These levels were comparable to those seen in OCE rats. We observed a significant decrease in DA concentrations in the ME and hyperprolactinemia in an exposure-dependent manner similar to that seen in OCE rats. It was concluded that chronic exposure to low levels of E(2) evokes oxidative stress in the AN to inhibit TIDA neuronal function, most probably leading to hyperprolactinemia.

Chronic estradiol-17ß exposure increases superoxide production in the rostral ventrolateral medulla and causes hypertension: reversal by resveratrol.

Women are exposed to estrogen in several forms, such as oral contraceptive pills and hormone replacement therapy. Although estrogen was believed to be cardioprotective, lately, its beneficial effects are being questioned. Recent studies indicate that oxidative stress in the rostral ventrolateral medulla (RVLM) may play a role in the development of hypertension. Therefore, we hypothesized that chronic exposure to low levels of estradiol-17ß (E(2)) leads to hypertension in adult-cycling female Sprague Dawley (SD) rats potentially through generation of superoxide in the RVLM. To test this hypothesis, young adult (3 or 4 mo old) female SD rats were either sham-implanted or implanted (subcutaneously) with slow-release E(2) pellets (20 ng/day) for 90 days. A group of control and E(2)-treated animals were fed lab chow or chow containing resveratrol (0.84 g/kg of chow), an antioxidant. Rats were implanted with telemeters to continuously monitor blood pressure (BP) and heart rate (HR). At the end of treatment, the RVLM was isolated for measurements of superoxide. E(2) treatment significantly increased mean arterial pressure (mmHg) and HR (beats/min) compared with sham rats (119.6 ± 0.8 vs. 105.1 ± 0.7 mmHg and 371.7 ± 1.5 vs. 354.4 ± 1.3 beats/min, respectively; P < 0.0001). Diastolic and systolic BP were significantly increased in E(2)-treated rats compared with control animals. Superoxide levels in the RVLM increased significantly in the E(2)-treated group (0.833 ± 0.11 nmol/min·mg) compared with control (0.532 ± 0.04 nmol/min·mg; P < 0.05). Treatment with resveratrol reversed the E(2)-induced increases in BP and superoxide levels in the RVLM. In conclusion, these findings support the hypothesis that chronic exposure to low levels of E(2) induces hypertension and increases superoxide levels in the RVLM and that this effect can be reversed by resveratrol treatment.

Prenatal exposure to bisphenols affects pregnancy outcomes and offspring development in rats.

The objective of this study was to evaluate the effects of gestational exposure to low doses of bisphenol A (BPA), bisphenol S (BPS), and bisphenol F (BPF) on pregnancy outcomes and offspring development. Pregnant Sprague-Dawley rats were orally dosed with vehicle, 5 µg/kg body weight (BW)/day of BPA, BPS and BPF, or 1 µg/kg BW/day of BPF on gestational days 6-21. Pregnancy and gestational outcomes, including number of abortions and stillbirths, were monitored. Male and female offspring were subjected to morphometry at birth, followed by pre- and post-weaning body weights, post-weaning food and water intakes, and adult organ weights. Ovarian follicular counts were also obtained from adult female offspring. We observed spontaneous abortions in over 80% of dams exposed to 5 µg/kg of BPF. BPA exposure increased Graafian follicles in female offspring, while BPS and BPF exposure decreased the number of corpora lutea, suggesting reduced ovulation rates. Moreover, BPA exposure increased male kidney and prostate gland weights, BPF decreased epididymal adipose tissue weights, and BPS had modest effects on male abdominal adipose tissue weights. Prenatal BPS exposure reduced anogenital distance (AGD) in male offspring, suggesting possible feminization, whereas both BPS and BPA induced oxidative stress in the testes. These results indicate that prenatal exposure to BPF affects pregnancy outcomes, BPS alters male AGD, and all three bisphenols alter certain organ weights in male offspring and ovarian function in female offspring. Altogether, it appears that prenatal exposure to BPA or its analogues can induce reproductive toxicity even at low doses.

Independent and combined effects of Bisphenol A and Diethylhexyl Phthalate on gestational outcomes and offspring development in Sprague-Dawley rats.

Bisphenol A (BPA) and Diethylhexyl Phthalate (DEHP) are well-studied endocrine disrupting chemicals (EDCs), however, the effects of mixtures of these EDCs are not. To assess the consequences of prenatal exposure to a mixture of these EDCs, dams were orally administered either saline (control), BPA (5 µg/kg BW/day), high dose DEHP (HD-D; 7.5 mg/kg BW/day), or a combination of BPA with HD-D in experiment 1; saline, BPA (5 µg/kg BW/day), low-dose DEHP (LD-D; 5 µg/kg BW/day) or a combination of BPA with LD-D in experiment 2. Gestational weights, number of abortions, litter size and weights, number of live births and stillbirths were recorded. Morphometric measures were obtained at birth and body weight, food and water intake were monitored weekly from postnatal weeks 3-12. Offspring were sacrificed at 16-24 weeks of age and organ weights were measured. The abortion rate of dams exposed to HD-D and the mixtures, BPA + LD-D and BPA + HD-D were higher at 9, 14 and 27% respectively. Prenatal exposure to BPA or HD-D significantly decreased relative thymus weights in male but not female offspring. Apoptotic cells were detected in thymus sections of both male and female offspring prenatally exposed to DEHP. Relative heart weights increased in BPA + HD-D exposed male offspring compared to the other groups. The results indicate that a mixture of BPA and DEHP, produced a pronounced effect on pregnancy outcomes. Male offspring appear to be more susceptible to the programming effects of these EDCs or their mixture suggesting a need to reconsider the possible additive, antagonistic or synergistic effects of EDC mixtures.

Oleic acid stimulates monoamine efflux through PPAR-α: Differential effects in diet-induced obesity.

Obesity continues to be a growing health concern around the world, and elevated levels of free fatty acids as a result of high-fat intake might play a role in neuroendocrine alterations leading to obesity. However, it is unclear how fatty acids affect neuroendocrine functions and energy metabolism. Since hypothalamic monoamines play a crucial role in regulating neuroendocrine functions relating to energy balance, we investigated the direct effects of oleic acid on hypothalamic monoamines and hypothesized that oleic acid would activate peroxisome proliferator-activated receptor alpha (PPAR-α), a nuclear transcription factor involved with fatty acid metabolism, to affect monoamines. We also hypothesized that this response would be subdued in diet-induced obesity (DIO). To test these hypotheses, hypothalami from Sprague Dawley and DIO rats were incubated with 0 (Control), 0.00132 mM, 0.132 mM, 1.32 mM oleic acid, 50 µM MK 886 (a selective PPAR- α antagonist), or oleic acid + MK 886 in Krebs Ringers Henseleit (KRH) solution. HPLC-EC was used to measure monoamine levels in perfusates. Oleic acid produced a significant increase in norepinephrine, dopamine, and serotonin levels in a dose-dependent manner, and incubation with MK886 blocked these effects. The effect of oleic acid on hypothalamic monoamines was attenuated in DIO rats. These findings suggest that PPARα probably plays an essential role in fatty acid sensing in the hypothalamus, by affecting monoamine efflux and DIO rats are resistant to the effects of oleic acid.

Leptin and HPA axis activity in diabetic rats: Effects of adrenergic agonists.

Type I Diabetes (T1D) is associated with reduced leptin levels and increased stress axis activity marked by elevations in norepinephrine (NE) levels in the paraventricular nucleus (PVN) of the hypothalamus. We hypothesized that leptin suppresses stress axis activity in T1D through central and peripheral mechanisms. In the first experiment, adult male Sprague Dawley rats were implanted with a cannula in the PVN and randomly divided into a non-diabetic group treated with vehicle (n = 6) and a diabetic group treated with streptozotocin (n = 13). Food intake and water intake was measured for 14 days. On the last day, a subset of diabetic rats were treated with 500 µg of leptin i.p. Rats were subjected to push-pull perfusion of the PVN and hourly blood sampling for 5 h. In the next experiment, diabetic rats were treated either with an alpha-2 adrenergic agonist, clonidine (CLON), or a beta adrenergic agonist isoproterenol (ISO), to reverse the effects of leptin. Rats were subjected to push pull perfusion and hourly blood sampling. In experiment 1, T1D increased food intake, water intake, NE release in the PVN and circulating CS levels. Leptin treatment decreased NE release modestly but produced a robust reduction in corticosterone (CS) levels. In experiment 2, CLON but not ISO was able to reverse the effect of leptin on NE levels in the PVN, however, both agonists were capable of blocking leptin's effects on circulating CS. These results suggest that in diabetic rats, the sensitivity of the hypothalamus to beta adrenergic agonists is altered, while the adrenals remain sensitive to both alpha and beta adrenergic agonists.

Chronic estrogen affects TIDA neurons through IL-1ß and NO: effects of aging.

Women are chronically exposed to estrogens through oral contraceptives, hormone replacement therapy or environmental estrogens. We hypothesized that chronic exposure to low levels of estradiol-17ß (E2) can induce inflammatory and degenerative changes in the tuberoinfundibular dopaminergic (TIDA) system leading to reduced dopamine synthesis and hyperprolactinemia. Young (Y; 3­4 months) and middle-aged (MA; 10­12 months) Sprague-Dawley rats that were intact or ovariectomized (OVX) were either sham-implanted or implanted with a slow-release E2 pellet (20 ng E2/day for 90 days). To get mechanistic insight, adult 3- to 4-month-old WT, inducible nitric oxide synthase (iNOS) and IL-1 receptor (IL-1R) knockout (KO) mice were subjected to a similar treatment. Hypothalamic areas corresponding to the TIDA system were analyzed. E2 treatment increased IL-1ß protein and nitrate levels in the arcuate nucleus of intact animals (Y and MA). Nitration of tyrosine hydroxylase in the median eminence increased with E2 treatment in both intact and OVX animals. There was no additional effect of age. This was accompanied by a reduction in dopamine levels and an increase in prolactin in intact animals. E2 treatment increased nitrate and reduced dopamine levels in the hypothalamus and increased serum prolactin in WT mice. In contrast, the effect of E2 on nitrate levels was blocked in IL-1R KO mice and the effect on dopamine and prolactin were blocked in iNOS KO animals. Taken together, these results show that chronic exposure to low levels of E2 decreases TIDA activity through a cytokine-nitric oxide-mediated pathway leading to hyperprolactinemia and that aging could promote these degenerative changes.

Responsiveness of hypothalamo-pituitary-adrenal axis to leptin is impaired in diet-induced obese rats.

BACKGROUND/OBJECTIVES: Diet-induced obese (DIO) rats have altered stress (HPA) axis activity compared to diet-resistant (DR) rats when chronically exposed to a high-fat (HF) diet. Since stress axis is tightly regulated by leptin, an adipocyte-secreted hormone that is important for controlling body weight, we hypothesized that leptin action is impaired in DIO rats leading to alterations in HPA axis activity. SUBJECTS/METHODS: We intraperitoneally injected selectively bred DIO and DR rats with either saline or recombinant rat leptin. HPA axis activity was assessed by measuring norepinephrine (NE) in the paraventricular nucleus (PVN), corticotropin-releasing hormone (CRH) in the median eminence, and serum corticosterone (CORT). To test if HF exposure duration and the corresponding increase in leptin differentially affects HPA axis activity, we placed animals on a chow or HF diet for 1 or 6 weeks. RESULTS: Leptin injection significantly increased serum leptin levels in both DIO and DR animals. It also reduced PVN NE in both groups, indicating that noradrenergic neurons in both groups remain responsive to leptin. HF diet duration-dependently increased serum leptin only in DIO animals whereas PVN NE increased in both groups. While DR rats responded to HF diet by increasing CRH and CORT at both time-points, responses in DIO rats varied, suggesting that they have altered HPA axis activity that may be dependent on HF-induced leptin levels and/or signaling. To understand the underlying mechanisms, we measured pSTAT-3, a marker of leptin signaling, in brainstem noradrenergic neurons and found reduced pSTAT-3 in A1 region of HF-fed DIO rats. We also found higher serum free fatty acids (FFAs) and a pro-inflammatory cytokine, IL-1ß. CONCLUSIONS: Collectively, these findings reveal that DIO rats have inherent neuroendocrine impairment in NE-HPA axis circuitry that worsens with the extent of HF diet exposure, possibly due to brainstem leptin resistance and/or elevated circulating FFAs and IL-1ß.

Systemic administration of leptin decreases plasma corticosterone levels: role of hypothalamic norepinephrine.

Leptin, an adipocyte-derived hormone, is known to regulate a variety of neuroendocrine functions. It inhibits the hypothalamo-pituitary-adrenal axis (HPA) in several animal models, however, the exact mechanism by which it does so is not known. Since norepinephrine (NE) is a key regulator of the HPA axis, we hypothesized that leptin could suppress HPA activity by decreasing NE levels. To study this, we implanted adult male Sprague-Dawley rats with both a push-pull cannula in the paraventricular nucleus (PVN) and a catheter in the jugular vein. Animals were treated with either 0 or 100 microg or 500 microg of recombinant rat leptin (Lep). Push-pull perfusion was performed from 1000-1600 h. Perfusate samples were collected every 30 min and analyzed for NE levels using HPLC-EC. Blood samples were collected every 60 min and analyzed for corticosterone (CS) levels. To further understand the role of NE in this phenomenon animals were treated with either an alpha1-adrenergic agonist, phenylephrine (PHE; 0.5 mg/kg BW), an alpha2-adrenergic agonist, clonidine (CLON; 0.6 mg/kg BW), or a beta-adrenergic agonist, isoproterenol (ISO; 0.2 mg/kg BW) alone or in combination with 500 microg of Lep. Pre-treatment and hourly post-treatment blood samples were collected, plasma was separated and analyzed for CS levels. Leptin administration decreased NE release in the PVN significantly by 30 min (p<0.05). It also significantly reduced plasma CS levels at 240 and 300 min (p<0.05). Administration of either PHE or CLON in combination with leptin prevented the leptin-induced decrease in CS. In contrast, administration of ISO along with leptin did not prevent the leptin-induced decrease in CS. These results indicate that leptin decreases hypothalamic NE and plasma CS and that this effect is most probably mediated through alpha-adrenergic receptors.

Particulate matter, oxidative stress and neurotoxicity.

Particulate matter (PM), a component of air pollution has been epidemiologically associated with sudden deaths, cardiovascular and respiratory illnesses. The effects are more pronounced in patients with pre-existing conditions such as asthma, diabetes or obstructive pulmonary disorders. Clinical and experimental studies have historically focused on the cardiopulmonary effects of PM. However, since PM particles carry numerous biocontaminants that are capable of triggering free radical production and cytokine release, the possibility that PM may affect organs systems sensitive to oxidative stress must be considered. Four independent studies that summarize the neurochemical and neuropathological changes found in the brains of PM exposed animals are described here. These were recently presented at two 2007 symposia sponsored by the Society of Toxicology (Charlotte, NC) and the International Neurotoxicology Association (Monterey, CA).

Chronic exposure to low doses of estradiol-17ß increases blood pressure in young female rats: A possible role for central Endothelin-1.

Previously, we demonstrated that chronic exposure to low levels of estradiol-17ß (E2) increases mean arterial pressure (MAP) in young female Sprague-Dawley (SD) rats, however, the underlying mechanisms are unclear. Since endothelin-1 (ET-1) is implicated in blood pressure (BP) regulation, we hypothesized that E2's effects on MAP are mediated through central ET-1. To test this, young female SD rats were either sham implanted or implanted s.c. with slow-release E2 pellets (20 ng/day for 90 days). BP was monitored by telemetry. After 75 days of E2 exposure, ETA antagonist or vehicle was administered i.c.v. After 90 days of E2 exposure, rats were sacrificed, and the paraventricular nucleus (PVN) and rostral ventrolateral medulla (RVLM) were microdissected for gene expression and protein analysis of ET-1 and its receptors. E2 exposure increased MAP after pellet implantation. Gene expression of ET-1 and ETA but not ETB receptors were upregulated in the PVN and RVLM of E2 treated animals. Further, the protein levels of ETA receptor were also increased in the PVN of E2 treated animals. However, i.c.v. infusion of the ETA antagonist did not completely block the increase in blood pressure. Our results suggest that increases in central ET-1 activity could possibly play a role in chronic E2-induced increase in BP but further studies are needed to completely understand the contribution of ET-1 in this phenomenon.

Activation of the stress axis and neurochemical alterations in specific brain areas by concentrated ambient particle exposure with concomitant allergic airway disease.

OBJECTIVE: Exposure to ambient particulate matter (PM) has been linked to respiratory diseases in people living in urban communities. The mechanism by which PM produces these diseases is not clear. We hypothesized that PM could act on the brain directly to stimulate the stress axis and predispose individuals to these diseases. The purpose of this study was to test if exposure to PM can affect brain areas involved in the regulation of neuroendocrine functions, especially the stress axis, and to study whether the presence of preexisting allergic airway disease aggravates the stress response. DESIGN: Adult male rats (n = 8/group) with or without ovalbumin (OVA)-induced allergic airway disease were exposed to concentrated air particles containing PM with an aerodynamic diameter pound 2.5 microm (PM(2.5)) for 8 hr, generated from ambient air in an urban Grand Rapids, Michigan, community using a mobile air research laboratory (AirCARE 1). Control animals were exposed to normal air and were treated with saline. MEASUREMENTS: A day after PM(2.5) exposure, animals were sacrificed and the brains were removed, frozen, and sectioned. The paraventricular nucleus (PVN) and other brain nuclei were microdissected, and the concentrations of aminergic neurotransmitters and their metabolites were measured using high-performance liquid chromatography with electrochemical detection. Serum corticosterone levels were measured using radioimmunoassay. RESULTS: A significant increase in the concentration (mean +/- SE, pg/microg protein) of norepinephrine in the PVN was produced by exposure to concentrated ambient particles (CAPs) or OVA alone (12.45 +/- 2.7 and 15.84 +/- 2.8, respectively) or after sensitization with OVA (19.06 +/- 3.8) compared with controls (7.98 +/- 1.3 ; p < 0.05). Serum corticosterone (mean +/- SE, ng/mL) was significantly elevated in the OVA + CAPs group (242.786 +/- 33.315) and in the OVA-presensitized group (242.786 +/- 33.315) compared with CAP exposure alone (114.55 +/- 20.9). Exposure to CAPs (alone or in combination with OVA pretreatment) can activate the stress axis, and this could probably play a role in aggravating allergic airway disease.

Systemic Interleukin-1beta stimulates the simultaneous release of norepinephrine in the paraventricular nucleus and the median eminence.

Interleukin-1beta (IL-1beta), a cytokine with pronounced central effects such as fever, anorexia, analgesia, etc., is also known to activate the hypothalamo-pituitary-adrenal (HPA) axis. Corticotropin releasing hormone (CRH) neurons located in the hypothalamus are important for HPA activation. The cell bodies of CRH neurons are located in the paraventricular nucleus (PVN) and their terminals are present in the median eminence (ME). Although the catecholamines, norepinephrine (NE) and dopamine (DA) are believed to be crucial factors in the stimulation of CRH neurons, it is not clear if they affect the cell bodies or terminals of these neurons to cause HPA activation. This study was done to determine if IL-1beta affects NE and DA release at the level of CRH cell bodies or their terminals. Adult male Sprague-Dawley rats were implanted with two push-pull cannulae, one in the PVN and another in the ME, and were subjected to push-pull perfusion. They were treated either with 0, 1 or 5 microg of IL-1beta. Perfusates were collected for 2 h after treatment and analyzed for NE concentrations using HPLC-EC. NE levels in the control and low dose groups did not change significantly during the entire period of observation both in the PVN and ME. In contrast, treatment with 5 microg of IL-1beta produced a marked increase in NE release in the PVN at 20 and 40 min post-treatment. NE release in the ME increased from 10 to 140 min post-treatment. There were no significant changes in the release of DA from both these areas. These results indicate that IL-1beta increases NE levels both in the PVN and in the ME and this could be a possible mechanism by which it stimulates the HPA axis.