VANL-100 Attenuates Beta-Amyloid-Induced Toxicity in SH-SY5Y Cells.

Antioxidants are being explored as novel therapeutics for the treatment of neurodegenerative diseases such as Alzheimer's disease (AD) through strategies such as chemically linking antioxidants to synthesize novel co-drugs. The main objective of this study was to assess the cytoprotective effects of the novel antioxidant compound VANL-100 in a cellular model of beta-amyloid (Aß)-induced toxicity. The cytotoxic effects of Aß in the presence and absence of all antioxidant compounds were measured using the 3-(4,5-dimethylthiazol-2-yl)2-5-diphenyl-2H-tetrazolium bromide (MTT) assay in SH-SY5Y cells in both pre-treatment and co-treatment experiments. In pre-treatment experiments, VANL-100, or one of its parent compounds, naringenin (NAR), alpha-lipoic acid (ALA), or naringenin + alpha-lipoic acid (NAR + ALA), was administrated 24 h prior to an additional 24-h incubation with 20 µM non-fibril or fibril Aß25-35. Co-treatment experiments consisted of simultaneous treatment with Aß and antioxidants. Pre-treatment and co-treatment with VANL-100 significantly attenuated Aß-induced cell death. There were no significant differences between the protective effects of VANL-100, NAR, ALA, and NAR + ALA with either form of Aß, or in the effect of VANL-100 between 24-h pre-treatment and co-treatment. These results demonstrate that the novel co-drug VANL-100 is capable of eliciting cytoprotective effects against Aß-induced toxicity.

Influence of staple line number and configuration on the leakage of small intestinal functional end-to-end stapled anastomosis: An ex vivo study.

OBJECTIVE: To determine the influence of the staple line configuration on the leakage of small intestinal functional end-to-end stapled anastomosis (FEESA). STUDY DESIGN: Experimental, ex vivo, randomized study. SAMPLE POPULATION: Jejunal segments (N = 72) from 10 mature, canine cadavers. METHODS: Jejunal segments (10 cm) were randomly assigned to a control group (8 segments) and 4 FEESA groups (16 segments/group (8 constructs/group)), according to the number of rows of staples used in the vertical (V) and transverse lines (T), respectively: Control, 2-row V/2-row T (2V/2T), 2-row V/3-row T (2V/3T), 3-row V/2-row T (3V/2T), 3-row V/3-row T (3V/3T). Initial leak pressure (ILP), maximum intraluminal pressure (MIP), and initial leakage location (ILL) were compared. RESULTS: The ILP (mean ± SD) for control segments, 2V/2T, 2V/3T, 3V/2T and 3V/3T were 321.38 ± 34.59, 32.88 ± 7.36, 50.13 ± 10.46, 34.38 ± 11.78, 69.88 ± 21.23 mmHg, respectively. All FEESAs initially leaked at lower pressures than intact segments. The only other differences detected between groups consisted of ILPs that were higher when FEESAs were closed with 3V/3T (69.88 ± 21.23 mmHg) than 2V/2T (32.88 ± 7.36, P < .001). Initial leakage occurred predominantly from the transverse staple line rather than the anastomotic crotch (P < .001). CONCLUSION: Placing 3 rows of staples in the transverse line (with or without a third row in the vertical staple line) improved resistance to leakage of FEESAs in normal cadaveric specimens. CLINICAL SIGNIFICANCE: The addition of a third row of staples in the transverse line (with or without a third row in the vertical staple line) in FEESAs should be further investigated as a strategy to reduce intestinal leakage clinically.

Evaluation of the performance of an endoscopic 3-mm electrothermal bipolar vessel sealing device intended for single use after multiple use-and-resterilization cycles.

OBJECTIVE: To evaluate the performance of an endoscopic 3-mm electrothermal bipolar vessel sealing device (EBVS) intended for single use after multiple use-and-resterilization cycles. STUDY DESIGN: Ex vivo study. SAMPLE POPULATION: Eight 3-mm EBVS handpieces. METHODS: Handpieces were subjected to a maximum of 15 cycles of testing, including simulated surgery, sealing and burst pressure testing of porcine carotid arteries, reprocessing, and hydrogen peroxide plasma resterilization. Failure was defined as two sequential vascular seal leakage events occurring at <250 mm Hg. Histological evaluation, maximum external temperature of the jaws, sealing time, tissue adherence, jaw surface characterization, and mechanical deterioration were studied. Failure rate was analyzed by using a Kaplan-Meier curve. Linear and ordinal logistic mixed models were used to analyze sealing time, handpiece jaw temperature, and adherence score. RESULTS: Mean ± SD diameter of arteries was 3.22 ± 0.35 mm. Failure was observed starting at cycle 10 and going up to cycle 13 in 37.5% (3/8) of the handpieces. Tissue adherence increased after each cycle (P < .001). Maximum external temperature (79.8°C ± 13.9°C) and sealing time (1.8 ± 0.5 seconds) were not significantly different throughout cycles up to failure. A flatter surface and large scratches were observed microscopically throughout the jaw surface after repeated use and resterilization. CONCLUSION: The 3-mm EBVS handpiece evaluated in this study can be considered safe to use for up to nine reuse-and-resterilization cycles. CLINICAL SIGNIFICANCE: These data provide the basis for establishing preliminary guidelines for the reuse and hydrogen peroxide plasma resterilization of an endoscopic 3-mm EBVS handpiece.

A co-drug conjugate of naringenin and lipoic acid mediates neuroprotection in a rat model of oxidative stress.

Using our in vitro and in vivo models of oxidative stress, the current study was designed to determine the neuroprotective potential of naringenin, alone or in combination with lipoic acid. In our mixed neuronal culture exposed to hypoxia and subsequent reoxygenation, naringenin was shown to provide significant neuroprotection against cell death at a concentration of 2.5 µmol/L. Lipoic acid (LA) did not produce neuroprotection at any concentration tested (0.25-100 µmol/L). In contrast, when naringenin was covalently combined with LA, producing a novel compound named "VANL-100", significant neuroprotection was observed at a concentration as low as 2×10-2  µmol/L (100-fold more potent). An ELISA for antioxidant capacity demonstrated that naringenin and VANL-100 likely resulted in neuroprotection by increasing the free radical scavenging capacity of the neuronal cells. Pretreatment of rats with the above compounds prior to middle cerebral artery occlusion (MCAO) followed by reperfusion, showed similar results. Naringenin significantly reduced infarct volume at a dose of 10 mg/kg while VANL-100 produced significant neuroprotection at a dose as low as 1×10-4  mg/kg (10 000-fold more potent). This VANL-100-induced neuroprotection persisted even when administered 1 and 3 hours into the reperfusion time course. Taken together, these results suggest that our novel compound, VANL-100 is neuroprotective, likely via a mechanism that involves increasing the antioxidant capacity of neuronal cells. Our results also show that VANL-100 is 100-10 000-fold more potent than the parent compounds, which adds to the growing evidence in support of combination therapy targeting oxidative stress in neurodegenerative diseases.

A novel synthetic chemical entity (UPEI-800) is neuroprotective in vitro and in an in vivo rat model of oxidative stress.

In this study, we tested a novel synthetic pyrazole-containing compound, 5-amino-1-phenyl-1H-pyrazole-4-carbonitrile (APPC), as an antioxidant in both in vitro and in vivo models of oxidative stress. In addition, the utility of covalently combining APPC with another well-established antioxidant, lipoic acid (LA), was also tested in both models. The in vitro results demonstrated that pretreatment with APPC in a mixed neuronal-glial culture exposed to oxygen-glucose deprivation (OGD) followed by reoxygenation-refeeding, resulted in significant neuroprotection at concentrations between 2.5 to 25 µmol/L. In contrast, LA was not neuroprotective following OGD alone or following reoxygenation-refeeding. However, the synthetic covalent combination of APPC with LA, named "UPEI-800", resulted in significant neuroprotection at concentrations between 0.027 and 2.7 µmol/L (100-fold more potent than APPC alone), an effect shown to be correlated with increased cellular antioxidant capacity. Further, in an in vivo model of ischaemia-reperfusion injury following transient occlusion of the middle cerebral artery (tMCAO), both APPC (0.1 and 1.0 mg/kg) and UPEI-800 (1×10-3  mg/kg) provided significant neuroprotection. Consistent with the in vitro findings, the in vivo results following tMCAO also demonstrated a 100-fold increase in the potency of the covalently linked compound UPEI-800 compared to APPC alone.

Guanosine protects against reperfusion injury in rat brains after ischemic stroke.

After ischemic stroke, early thrombolytic therapy to reestablish tissue perfusion improves outcome but triggers a cascade of deleterious cellular and molecular events. Using a collaborative approach, our groups examined the effects of guanosine (Guo) in response to ischemic reperfusion injury in vitro and in vivo. In a transient middle cerebral artery occlusion (MCAO) in rats, Guo significantly reduced infarct volume in a dose-dependent manner when given systemically either immediately before or 30 min, but not 60 min, after the onset of the 5.5-hr reperfusion period. In a separate experiment, Guo significantly reduced infarct volume after 24 hr of reperfusion when administered 5 min before reperfusion. Western blot analysis did not reveal any significant changes either in endoplasmic reticulum (ER) stress proteins (GRP 78 and 94) or HSP 70 or in levels of m-calpain. In vitro oxygen and glucose deprivation (OGD) significantly increased production of both reactive oxygen species (ROS) and interleukin-8 (IL-8) in the primary astrocytes. Guo did not alter ROS or IL-8 production when given to the astrocytes before OGD. However, Guo when added to the cells prior to or 30 min after reperfusion significantly reduced IL-8 release but not ROS formation. Our study revealed a dose- and time-dependent protective effect of Guo on reperfusion injury in vitro and vivo. The mechanisms by which Guo exerts its effect are independent of unfolded proteins in ER or the level of intracellular calcium or ROS formation. However, the effect may be induced, at least partially, by inhibiting IL-8, a marker of reperfusion-triggered proinflammatory events.

UPEI-100, a conjugate of lipoic acid and apocynin, mediates neuroprotection in a rat model of ischemia/reperfusion.

Previous work in our laboratory has provided evidence that preadministration of apocynin and lipoic acid at subthreshold levels for neuroprotection enhanced the neuroprotective capacity when injected in combination. Therefore, the present investigation was designed to determine whether a co-drug consisting of lipoic acid and apocynin functional groups bound by a covalent bond, named UPEI-100, is capable of similar efficacy using a rodent model of stroke. Male rats were anesthetized with Inactin (100 mg/kg iv), and the middle cerebral artery was occluded for 6 h or allowed to reperfuse for 5.5 h following a 30-min occlusion (ischemia/reperfusion, I/R). Preadministration of UPEI-100 dose-dependently decreased infarct volume in the I/R model (P < 0.05), but not in the middle cerebral artery occlusion model of stroke. Using the optimal dose, we then injected UPEI-100 during the stroke or at several time points during reperfusion, and significant neuroprotection was observed when UPEI-100 was administered up to 90 min following the start of reperfusion (P < 0.05). A time course for this neuroprotective effect showed that UPEI-100 resulted in a decrease in infarct volume following 2 h of reperfusion compared with vehicle. The time course of this neuroprotective effect was also used to study several mediators along the antioxidant pathway and showed that UPEI-100 increased the level of mitochondrial superoxide dismutase and oxidized glutathione and decreased a marker of lipid peroxidation due to oxidative stress (HNE-His adduct formation). Taken together, the data suggest that UPEI-100 may utilize similar pathways to those observed for the two parent compounds; however, it may also act through a different mechanism of action.

Naturally Occurring Antioxidant Therapy in Alzheimer's Disease.

It is estimated that the prevalence rate of Alzheimer's disease (AD) will double by the year 2040. Although currently available treatments help with symptom management, they do not prevent, delay the progression of, or cure the disease. Interestingly, a shared characteristic of AD and other neurodegenerative diseases and disorders is oxidative stress. Despite profound evidence supporting the role of oxidative stress in the pathogenesis and progression of AD, none of the currently available treatment options address oxidative stress. Recently, attention has been placed on the use of antioxidants to mitigate the effects of oxidative stress in the central nervous system. In preclinical studies utilizing cellular and animal models, natural antioxidants showed therapeutic promise when administered alone or in combination with other compounds. More recently, the concept of combination antioxidant therapy has been explored as a novel approach to preventing and treating neurodegenerative conditions that present with oxidative stress as a contributing factor. In this review, the relationship between oxidative stress and AD pathology and the neuroprotective role of natural antioxidants from natural sources are discussed. Additionally, the therapeutic potential of natural antioxidants as preventatives and/or treatment for AD is examined, with special attention paid to natural antioxidant combinations and conjugates that are currently being investigated in human clinical trials.

CYP17, catechol-o-methyltransferase, and glutathione transferase M1 genetic polymorphisms, lifestyle factors, and breast cancer risk in women on Prince Edward Island.

Genetic polymorphisms in enzymes controlling the formation and disposition of estrogens and their metabolites have been shown to influence breast cancer risk. Environmental and lifestyle factors may interact with estrogen metabolism polymorphisms to influence breast cancer risk. We studied the role of lifestyle factors and genetic polymorphisms in estrogen metabolism in women from Prince Edward Island (PEI), a small province of 135,000 people on the east coast of Canada. Women (207 cases; 621 controls) were matched on age, menopausal status, and family history of breast cancer. The predominant lifestyle risk factors previously reported to influence breast cancer risk such as body mass index (BMI), parity, and smoking had similar influences in the PEI population. Genetic polymorphisms in CYP17, GSTM1, and catechol-O-methyltransferase (COMT) were not associated with a general increase in breast cancer risk. However, the CYP17 A2/A2 genotype was only observed in women with estrogen receptor (ER) positive breast cancer and not in ER negative breast cancer. The increased risk associated with elevated BMI was only observed in women homozygous for the CYP17 and COMT reference alleles. Similarly, the increased risk associated with extended use of oral contraceptives (≥ 15years), was only observed in women homozygous for the reference alleles of CYP17 and COMT. The GSTM1 homozygous gene deletion was associated with a significantly increased risk of breast cancer in postmenopausal women with a family history of breast cancer risk. These results suggest the polymorphic genes that control estrogen formation and disposition interact significantly with other risk factors to influence breast cancer risk.

Substance P and cocaine employ convergent mechanisms to depress excitatory synaptic transmission in the rat nucleus accumbens in vitro.

Substance P (SP) has been reported to produce effects on excitatory synaptic transmission in the nucleus accumbens (NAc) that are similar to those induced by cocaine. To address the question of whether SP serves as an endogenous mediator producing cocaine-like effects that are known to be D1-receptor-mediated, we tested the hypothesis that the effects of SP and cocaine on excitatory postsynaptic currents (EPSCs) in the NAc occlude one another. We report here that SP and SP(5-11) actions occlude the effect of cocaine and vice versa. SP, SP(5-11) and cocaine all depressed evoked, non-N-methyl-D-aspartate (NMDA) receptor-mediated synaptic currents in a concentration-dependent manner, with EC50 values of 0.12, 0.17 and 8.3 microm, respectively. Although cocaine was the least potent, it was most efficacious. SP, SP(5-11) and cocaine all suppressed isolated NMDA receptor-mediated evoked EPSCs. SP(5-11) (1 microm)-induced EPSC depression was blocked by the neurokinin-1 antagonist L732138 and by the D1-like receptor antagonist SCH23390. Pretreatment of slices with cocaine (30 microm) depressed the EPSC by 39.1% +/- 4.8%. Application of SP or SP(5-11) (1 microm) at the peak of the cocaine depressive effect on the EPSC did not produce any additional diminution of the response (5.7% +/- 2.8%). In the reverse experiments, in which either SP or SP(5-11) was applied first, subsequent application of cocaine at the peak of the peptide's effect (30.3% +/- 2.3%) produced a further but smaller depression (15.5% +/- 3.6%) of the remaining EPSC. These data indicate that cocaine and SP produce similar effects on excitatory synaptic transmission in the NAc, and that their actions occlude one another. This suggests that SP may act like cocaine in its absence, and may be an endogenous trigger for the reward and behaviors associated with cocaine.

Estrogen may contribute to ischemic tolerance through modulation of cellular stress-related proteins.

Ischemic tolerance describes a phenomenon whereby subcritical stimuli evoke cellular protective mechanisms resulting in increased tolerance to subsequent ischemia. In the present study we propose that the cytoprotective effects attributed to 17beta-estradiol and tunicamycin in an in vivo rodent model of ischemia are reflected by changes in neuronal tissue levels of m-calpain, HSP70, GRP94 and GRP78. Rats pretreated with 17beta-estradiol, tunicamycin or both demonstrated dose-dependent reductions in infarct area following 4 h of permanent middle cerebral artery occlusion (MCAO). Western blot analysis revealed that 4 h of MCAO was associated with decreased cortical expression of HSP70 and m-calpain and increased expression of GRP78. Pretreatment with 12.5 microg/kg 17beta-estradiol did not change this pattern of protein expression following MCAO. While GRP94 expression was elevated in sham-operated rats pretreated with 17beta-estradiol, the ensuing ischemic tolerance did not appear to be mediated by changes in cellular stress proteins. Pretreatment with 50 microg/kg tunicamycin significantly reduced HSP70 in cortical tissue samples taken from sham-operated rats and appeared to attenuate the threshold for activation of m-calpain in rats undergoing 4 h of MCAO. Lastly, a combined treatment in which rats undergoing MCAO were pretreated with both tunicamycin (24 h prior) and 17beta-estradiol (30 min prior) was associated with an attenuated stress response as indicated by reduced expression of GRP78 and GRP94 when compared to saline-treated controls. The results of this study suggest that the ischemic tolerance observed following MCAO in rats pretreated with either 17beta-estradiol or tunicamycin is likely mediated in part through differential effects on cellular stress proteins.

Incorporation of nanoparticles into transplantable decellularized matrices: Applications and challenges.

Decellularization of tissues can significantly improve regenerative medicine and tissue engineering by producing natural, less immunogenic, three-dimensional, acellular matrices with high biological activity for transplantation. Decellularized matrices retain specific critical components of native tissues such as stem cell niche, various growth factors, and the ability to regenerate in vivo. However, recellularization and functionalization of these matrices remain limited, highlighting the need to improve the characteristics of decellularized matrices. Incorporating nanoparticles into decellularized tissues can overcome these limitations because nanoparticles possess unique properties such as multifunctionality and can modify the surface of decellularized matrices with additional growth factors, which can be loaded onto the nanoparticles. Therefore, in this minireview, we highlight the various approaches used to improve decellularized matrices with incorporation of nanoparticles and the challenges present in these applications.

UPEI-400, a conjugate of lipoic acid and scopoletin, mediates neuroprotection in a rat model of ischemia/reperfusion.

Previously, our laboratory provided evidence that lipoic acid (LA) covalently bonded to various antioxidants, resulted in enhanced neuroprotection compared to LA on its own. The naturally occurring compound scopoletin, a coumarin derivative, has been shown in various in vitro studies to have both antioxidant and anti-inflammatory mechanism of actions. The present investigation was designed to determine if scopoletin on its own, or a co-drug consisting of LA and scopoletin covalently bonded together, named UPEI-400, would be capable of demonstrating a similar neuroprotective efficacy. Using a rat stroke model, male rats were anesthetized (Inactin®; 100 mg/kg, iv), the middle cerebral artery was permanently occluded for 6 h (pMCAO), or in separate animals, occluded for 30 min followed by 5.5 h of reperfusion (ischemia/reperfusion; I/R). Pre-administration of either scopoletin or UPEI-400 significantly decreased infarct volume in the I/R model (p < 0.05), but not in the pMCAO model of stroke. UPEI-400 was ∼1000 times more potent compared to scopoletin alone. Since UPEI-400 was only effective in a model of I/R, it is possible that it may act to enhance neuronal antioxidant capacity and/or upregulate anti-inflammatory pathways to prevent the neuronal cell death.

Role of polymorphic human cytochrome P450 enzymes in estrone oxidation.

Estrogen and its metabolites are believed to play important roles in breast cancer. The influence of genetic polymorphisms in the enzymes responsible for formation and disposition of estrogen on breast cancer risk may shed light on the importance of estrogen metabolites in this disease. However, for such studies to be valid, it is important to correctly identify the enzymes involved in estrogen bioactivation. Therefore, we assessed the human cytochrome P450-dependent oxidation of estrone using substrate concentrations that more closely approximate the maximum expected concentrations in breast tissue. The in vitro metabolism of estrone by recombinant human cytochrome P450 enzymes and human liver microsomes was studied. The formation of estrone metabolites (2-hydroxyestrone, 4-hydroxyestrone, and 16alpha-hydroxyestrone) was monitored by high-performance liquid chromatography. 2-Hydroxyestrone formation was catalyzed predominantly by CYP1A2, CYP1A1, and CYP1B1 enzymes; 4-hydroxyestrone formation was catalyzed predominantly by CYP1B1, CYP1A2, and CYP1A1 enzymes; and 16alpha-hydroxyestrone formation was catalyzed predominantly by CYP2C19, CYP1A1, and CYP3A5. This study confirms the important role of members of the CYP1 family in the 2-hydroxylation and 4-hydroxylation of estrone, but the enzymes identified as responsible for the 16alpha-hydroxylation of estrone are different from those previously identified. The relative importance of these enzymes in vivo would depend on the specific tissue expression of the enzymes. These enzymes are all known to be genetically variant in the human population, and additional studies to assess the role CYP1A2, CYP2C19, and CYP3A5 in breast cancer risk are indicated.

Estrogen reduces the severity of autonomic dysfunction in spinal cord-injured male mice.

Autonomic dysreflexia is an autonomic behavioural condition that manifests after spinal cord injury (SCI) and is characterized by acute, episodic hypertension following afferent stimulation below the level of the injury. Common triggers of autonomic dysreflexia include colorectal distension (CRD), and various somatic stimuli. The development of autonomic dysreflexia is dependent, in part, upon the degree of intraspinal inflammation and the resultant spinal neuroplastic changes that occur following SCI. 17beta-estradiol (E) has neuroprotective, anti-inflammatory and smooth muscle relaxant properties, and is therefore a candidate drug for the treatment and/or prevention of autonomic dysreflexia. Autonomic dysreflexia was assessed in adult male mice treated with E. We investigated whether E could be acting centrally by altering: (1) the size of the small diameter primary afferent arbor, (2) the degree of microglia/macrophage infiltration at the site of the injury, or (3) the amount of fibrous scarring present at the injury site. To determine whether E could be working through uncoupling protein-2 (UCP-2), a protein involved with inflammation and regulated by estrogen in some tissues, autonomic dysreflexia was assessed in E-treated adult male mice lacking UCP-2 (UCP-2 KO). 17beta-estradiol was equipotent at reducing autonomic dysreflexia in both UCP-2 KO and WT mice following CRD but not tail pinch. We have shown that E reduces autonomic dysreflexic responses to visceral but not somatic stimulation in male mice independent of the size of the primary afferent arbour, the degree of chronic inflammation, and the presence of UCP-2.

Relaxin-induced reduction of infarct size in male rats receiving MCAO is dependent on nitric oxide synthesis and not estrogenic mechanisms.

Relaxins are members of the insulin peptide superfamily. Previous evidence has shown that relaxin pretreatment reduces cortical infarct size in anesthetized, male rats receiving permanent middle cerebral artery occlusion (MCAO). Therefore, the current study was designed to determine if estrogenic mechanisms or nitric oxide production are involved in mediating this relaxin-induced neuroprotection. In separate groups of rats (n=4-6), the following drugs were injected directly into the cortex 30 min prior to MCAO: (a) relaxin, (b) relaxin and estrogen, and (c) relaxin and an estrogen receptor antagonist (ICI 182,780). To investigate the involvement of nitric oxide, relaxin or relaxin and an inhibitor of endothelial nitric oxide synthase (L-NIO) were injected i.v. 30 min prior to MCAO. Saline-treated rats (both intracortical (i.c.) and intravenously (i.v.)) served as controls. Brains were harvested 4h post stroke, coronally sectioned using a brain matrix and stained using 2,3,5-triphenoltetrazolium chloride (TTC). Digital photographs were taken of brain sections and the ratio comparing the area of the infarct to the area of the ipsilateral hemisphere was calculated. Mean ratios were compared using ANOVA and Tukey's test. Intracortical and intravenous relaxin pretreatment significantly reduced the infarct area in the cortex by 33.7 and 58.6%, respectively compared to saline-treated controls. This effect was not dependent on an interaction with estrogenic receptors as co-injection of relaxin and ICI 182,780 did not reverse this effect. However, inhibition of nitric oxide synthase significantly reduced the relaxin-mediated neuroprotection suggesting that relaxin may induce the endothelin-NOS cascade in cerebral vasculature causing vasodilation and improved perfusion of neural tissue.

Relaxin pretreatment decreases infarct size in male rats after middle cerebral artery occlusion.

We studied the possible neuroprotective action of relaxin in a rat stroke model. Relaxin (10 ng in 200 nL saline) or saline was injected into the secondary somatosensory cortex of anesthetized rats. Thirty minutes after treatment, the right middle cerebral artery was occluded, causing ischemic conditions. Brains were removed 4 hours after stroke, and 1-mm coronal sections were stained using 2-3-5-triphenoltetrazolium chloride. Digital photographs were taken of the sections, and the ratio of infarct area to ipsilateral hemispheric area was calculated. Relaxin treatment significantly (P < .05) reduced this ratio compared with that of saline-treated controls. Results suggest that relaxin may prevent ischemia-induced cell death.

Estrogen in the parabrachial nucleus attenuates the sympathoexcitation following MCAO in male rats.

Recent investigations have provided evidence to suggest systemic estrogen administration prevented or reversed the sympathoexcitation observed following middle cerebral artery occlusion (MCAO) in male rats. The present investigation sought to determine the role of estrogen injected directly into the parabrachial nucleus (PBN) on the MCAO-induced sympathoexcitation as well as the role of the rostral ventrolateral medulla (RVLM) in mediating the sympathoexcitatory response. Male Sprague-Dawley rats were anesthetized with sodium thiobutabarbitol (100 mg/kg) and were instrumented to continuously record blood pressure, heart rate and renal sympathetic nerve activity (RSNA). Following occlusion of the middle cerebral artery, there was a significant increase in RSNA (from 3.8 +/- 0.4 to 8.3 +/- 0.6 microV/s; P < 0.05) which was significantly attenuated by the prior bilateral injection of estrogen (0.5 microM in 200 nl) into the PBN. Pre-injection of lidocaine (5% in 200 nl) directly into the RVLM resulted in only a slight reduction in the magnitude of the MCAO-induced sympathoexcitation (P > 0.05). Extracellular electrophysiological recordings from RVLM neurons demonstrated that MCAO did not produce any significant change in neuronal activity over the experimental time course (P > 0.05). Also, bilateral injection of estrogen into the PBN prior to MCAO or sham conditions did not result in any significant change in RVLM neuronal activity. These results indicate that estrogen receptors in the PBN play a major role in modulating the sympathoexcitatory response from ischemic forebrain nuclei, and that the pathway from the PBN to sympathetic preganglionic nuclei may not involve a synapse in the RVLM.

Sympathoexcitatory effects of estrogen in the insular cortex are mediated by GABA.

The current investigation examined the effect of estrogen in the insular cortex (IC) on autonomic tone and cardiac baroreceptor reflex function and sought to determine if modulation of neurotransmission was responsible for mediating this effect. Experiments were performed in Inactin-anaesthetized, male Sprague-Dawley rats. Animals were instrumented to record blood pressure, heart rate, vagal parasympathetic and renal sympathetic nerve activities, as well as cardiac baroreflex sensitivity (BRS). Direct, bilateral injection of 17beta-estradiol (0.5 microM; 200 nl/side) into the IC resulted in a significant increase in sympathetic tone (from 10 +/- 4 to 24 +/- 3) with no significant change in blood pressure, heart rate, parasympathetic tone or BRS measured at 30 min post-injection. This estrogen-induced effect was completely blocked by the co-injection of estrogen with the estrogen receptor antagonist, ICI 182, 780 (20 microM; 200 nl/side). Co-injection of estrogen with a GABA(B), NMDA or non-NMDA receptor antagonists did not effect the estrogen-induced increase in sympathetic tone. Co-injection of a sub-threshold dose of estradiol (0.125 microM; 200 nl/side) with the GABA(A) receptor antagonist, (+)-bicuculline (0.025 microM; 200 nl/side), resulted in an additive response to increase sympathetic nerve activity. These results suggest that estrogen acts on estrogen receptors to modulate GABA(A)-receptor-mediated neurotransmission within the IC to modulate sympathetic tone.

A pharmacological study on Berberis vulgaris fruit extract.

Berberis vulgaris fruit (barberry) is known for its antiarrhythmic and sedative effects in Iranian traditional medicine. The effects of crude aqueous extract of barberry on rat arterial blood pressure and the contractile responses of isolated rat aortic rings and mesenteric bed to phenylephrine were investigated. We also examined effect of the extract on potassium currents recorded from cells in parabrachial nucleus and cerebellum rejoins of rat brain. Administration of the extract (0.05-1 mg/100 g body weight of rat) significantly reduced the mean arterial blood pressure and heart rate in anaesthetized normotensive and desoxycorticosteron acetate-induced hypertensive rats in a dose-dependent manner. Concentration-response curves for phenylephrine effects on isolated rat aortic rings and the isolated mesenteric beds in the presence of the extract were significantly shifted to the right. Application of the extract (1-50 microg/ml) shifted the activation threshold voltage to more negative potentials, leading to an enhancement in magnitude of the outward potassium current recorded from cells present in rat brain slices of parabrachial nucleus and cerebellum. This effect on potassium current may explain the sedative and neuroprotective effects of barberry. The present data support the hypothesis that the aqueous extract of barberry has beneficial effects on both cardiovascular and neural system suggesting a potential use for treatment of hypertension, tachycardia and some neuronal disorders, such as epilepsy and convulsion.