The cardiovascular responses to mu opioid agonist and antagonist in conscious normal and obese rats.

Beta-endorphin decreases blood pressure in normal rats but increases blood pressure in obese rats. Since beta-endorphins can bind both mu opioid and kappa-opioid receptors we investigated the effect of a mu specific receptor agonist, D-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin (DAMGO) and a mu specific antagonist, D-Phe-Cys-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) on cardiovascular responses in conscious control and obese rats. Rats were also implanted with telemetry transmitters and intracerebroventricular (ICV) cannulas for recording and peptide administration. The mu agonist, DAMGO, increased blood pressure (BP) in control rats. DAMGO also increased BP in obese rats but only at high concentrations. The heart rate responses paralleled the MAP responses. CTAP, the mu antagonist, paradoxically increased the MAP in both control and obese rats. The responsiveness to the mu agonist and antagonist was greater in controls. In other animals the brains were excised and the ventral medial hypothalamic area removed and mu receptor expression determined using PCR. The expression of mu opioid receptors was increased in obese rats. We conclude that the mu opioids can stimulate cardiovascular responses, but the excitatory responsiveness was not increased in conscious obese rats.

Cerebral ischemia-induced apoptosis and necrosis in normal and diabetic rats: effects of insulin and C-peptide.

Neuronal apoptosis has been demonstrated to be a significant factor in neurological deficiencies associated with diabetes, and these deficiencies are exaggerated following ischemia. Diabetic rats have an increased basal level of apoptosis compared to non-diabetics and it has been previously demonstrated that infarct volumes were greater in diabetic animals following middle cerebral artery occlusion (MCAO) when compared to non-diabetics. In this study, we evaluated both the acute and chronic effects of insulin and/or C-peptide on CNS necrosis and apoptosis in non-diabetic and streptozotocin-induced diabetic rats following MCAO with reperfusion. Two brain areas, the sensori-motor cortex (layers-5 and 6) and the CA1 and CA3 sectors (pyramidal cell layers) of the hippocampus, were analyzed for apoptosis using TUNEL and Caspase-3 immunoreactivity. The chronic administration of a low maintenance concentration of insulin (2 U/kg), or the acute administration of insulin (2 U/kg) with or without C-peptide, did not alter the lesion volume or basal levels of apoptosis or the apoptotic levels in animals subjected to 2-h MCAO followed by 24-h reperfusion. However, both the acute or chronic administration of a high concentration of insulin (12 U/kg) significantly decreased lesion volume and apoptosis subsequent to 2-h MCAO followed by 24-h reperfusion. High dose insulin treatment also decreased the basal level of apoptosis. We conclude that in diabetic rats subjected to ischemia and reperfusion chronic insulin treatment decreased the basal apoptotic level, and both acute and chronic insulin decreased the MCAO-induced lesion volume and apoptosis. Maintenance insulin concentrations with or without C-peptide were without effect.

Cerebral ischemia induced apoptosis and necrosis in normal and diabetic rats.

Stroke is the third leading cause of death and disability, and the risk for ischemic stroke is greater in diabetics. Previous studies have demonstrated both structural and functional nervous system changes in diabetes, and these changes may be enhanced by apoptosis. In the present study, we evaluated several indexes of both necrosis and apoptosis in the CNS of normals and two different models of diabetes (insulinopenic and insulin-resistant). Studies were conducted following middle cerebral artery occlusion (MCAO) with or without reperfusion. The sensory motor cortex (layer-5 and -6) and the CA1 and CA3 sectors of the hippocampus were analyzed following MCAO. We observed that both insulinopenic and insulin-resistant diabetic rats have increased basal level of apoptosis that is uniformly and bilaterally distributed as indicated by both caspase-3 activity and TUNEL staining. Twenty-four hours after MCAO, apoptosis was further increased in both diabetic models. Reperfusion after a 2 h MCAO compared to 24 h MCAO was associated with a decrease in TUNEL staining and caspase-3 activity in the control animal but exacerbated apoptosis, especially in the hippocampus of insulin-resistant diabetic rats. MCAO-induced lesion volumes were greater in insulinopenic rats compared to insulin-resistant and control rats. We conclude that both insulinopenic and insulin-resistant diabetic animals have increased apoptosis in the CNS in response to MCAO, and restoration of blood flow especially in the insulinopenic diabetic animals paradoxically exacerbates this process. Furthermore, restoration of blood flow did not decrease lesion volume in insulinopenic diabetic animals.

A role for the central histaminergic system in the leptin-mediated increase in cardiovascular dynamics.

The central nervous system (CNS) histaminergic neurons have been shown to regulate feeding behavior and are a target of leptin in the brain. The present study aimed to examine the involvement of the histaminergic system in the leptin-mediated regulation of cardiovascular dynamics. We investigated the cardiovascular responses to the CNS administration of histamine, leptin and alpha-melanocyte stimulating hormone (alpha-MSH) both in the presence and absence of the histamine H1 antagonist, chlorpheniramine. The intracerebroventricular (i.c.v.) administration of histamine resulted in an immediate increase in both mean arterial pressure (MAP) and heart rate (HR) and vasoconstricted the iliac, renal and superior mesenteric vessels. The i.c.v. pretreatment with chlorpheniramine attenuated the histamine-induced increase in MAP, HR and decreased vascular conductance. The i.c.v. administration of leptin increased MAP and HR and decreased vascular conductance. The i.c.v. pretreatment with chlorpheniramine decreased the leptin-induced increase in MAP and the leptin-mediated iliac vasoconstriction. The i.c.v. administration of alpha-MSH also increased MAP, HR and decreased vascular conductance. However, pretreatment with chlorpheniramine did not influence the central alpha-MSH-mediated increase in MAP, HR and decreased vascular conductance. These results indicate that the central histaminergic system mediated by H1 receptors have a role in the central signaling pathway and is involved in leptin's regulation of cardiovascular dynamics. It appears that leptin directly or indirectly stimulates histaminergic neurons that lead to increased cardiovascular activity.

The effects of acute and chronic alpha melanocyte stimulating hormone (alphaMSH) on cardiovascular dynamics in conscious rats.

Alpha melanocyte stimulating hormone (alphaMSH) has been demonstrated to have regulatory functions in the periphery and central nervous system (CNS). alphaMSH plays a central role in the regulation of metabolic balance such as decreasing food intake, increasing sympathetic outflow and hypothalamic/pituitary function. Our laboratory has investigated the actions of alphaMSH on sympathetic and cardiovascular dynamics using anesthetized animals. In this study we determined both the acute and chronic effects of alphaMSH on cardiovascular and metabolic dynamics in conscious unrestrained rats. Animals were each implanted with a radio-telemetry transmitter for recording of cardiovascular parameters and subsequently instrumented with intracerebroventricular (ICV) cannulas. The acute ICV administration of alphaMSH significantly increased the mean arterial pressure (MAP) and heart rate (HR) when compared to artificial cerebrospinal fluid (ACSF) controls. On the other hand chronic alphaMSH infusion resulted in an initial increase in MAP and HR lasting for 2 days followed by a decrease in MAP. Chronic alphaMSH administration decreased physical activity and food intake but not weight gain. We conclude that in the conscious unrestrained animal the acute administration of alphaMSH increased MAP and HR, however, chronic infusion is associated with decreased MAP, physical activity and food intake.

Cardiovascular responses to central administration of mu and kappa opioid receptor agonist and antagonist in normal rats.

The response to centrally administered beta-endorphin has been characterized by decreasing sympathetic nervous activity and decreased cardiovascular tone. We investigated the effect of the central administration of both mu and kappa opioid receptor agonist and antagonists on cardiovascular responses. The administration of the mu agonist, DAMGO (0.2nmol) increased the mean arterial pressure (MAP) and stimulated iliac vasoconstriction while higher doses (2 and 20nmol) decreased MAP and stimulated iliac vasodilation. The administration of the kappa receptor agonist, Dynorphin decreased the MAP and stimulated superior mesenteric vasodilation. beta-Funaltrexamine reduced MAP and superior mesenteric vasodilation while nor-binaltorphimine increased MAP and iliac and superior mesenteric vasoconstriction. We conclude that mu receptor activation decrease or increase MAP depending on the mu agonist concentration. However, kappa receptor activation is consistently associated with a decrease in MAP.

The effect of CNS opioid on autonomic nervous and cardiovascular responses in diet-induced obese rats.

The intracerebroventricular (i.c.v.) infusion of beta-endorphin can cause either a decrease in blood pressure in normal rats or an increase in obese rats. Diet-induced obesity is associated with an increase of hypothalamic mu opioid receptors. Since beta-endorphins act by opioid receptors, we investigated the effect of CNS mu as well as kappa opioid receptor agonist and antagonist on mean blood pressure (MAP), heart rate (HR) and renal sympathetic nerve activity (RSNA) in male Wistar rats fed either a high fat (HF) (40% fat by weight) or a regular low fat (control) (4% fat by weight) diet. After a 12-week-feeding period the animals were implanted with i.c.v. cannulas and 3-5 days later they were anesthetized and instrumented to record MAP, HR and RSNA. HF rats have higher MAP and the i.c.v. injection of a mu opioid agonist (DAMGO) initially decreased the MAP and then increased MAP, HR and RSNA in the normal animals. The increase was greater in HF animals. The i.c.v. injection of the mu antagonist (beta-FNA) resulted in a significantly greater decrease in MAP in HF animals. beta-FNA increased the RSNA in the HF rats but decreased it in the normal rats. The kappa agonist (dynorphin) decreased MAP in normal rats followed by a return to baseline, but not in HF rats. The kappa antagonist, nor-binaltorphimine (N-BP), increased MAP and RSNA in normal rats and to a lesser extent in HF rats. These findings suggest that rats given a high fat diet have higher blood pressures and a greater mu opioid-mediated responsiveness with a greater mu opioid-mediated autonomic tone. Additionally there is a decreased kappa responsiveness and tone in the HF rats. Both these changes, increased mu and decreased kappa responsiveness could strongly contribute to the increased blood pressure in obese animals.

Insulin-mediated increase in sympathetic nerve activity is attenuated by C-peptide in diabetic rats.

Connecting peptide (C-peptide) is secreted along with insulin in equimolar amounts into portal circulation in response to beta cell stimulation. The biological function of C-peptide had been mostly limited to establishing the secondary and tertiary structure of proinsulin. Recent studies have suggested that C-peptide can impact several functions, such as autonomic and sensory nerve function, insulin secretion, and microvascular blood flow. In this study we examined the effects of C-peptide in the presence or absence of insulin on cardiovascular and sympathetic nerve activity in both normal and streptozotocin (STZ)-induced diabetic Wistar rats. Animals were made diabetic by a single intravenous injection of STZ (50 mg/kg) and maintained for 6 weeks. The diabetic animals had higher plasma glucose, lower plasma insulin, and C-peptide, compared with the normal animals. To characterize cardiovascular and autonomic nervous responses, the animals were anesthetized with urethane/alpha-chloralose and instrumented for the recording of mean arterial pressure (MAP), heart rate (HR), and lumbar sympathetic nerve activity (LSNA). A bolus administration of C-peptide alone did not alter MAP, HR, or LSNA in normal or diabetic animals. The bolus administration of insulin alone increased HR and LSNA in normal and diabetic animals. However, the administration of insulin plus C-peptide attenuated the increase in HR in normals and the increase in LSNA in diabetic rats. We concluded that the C-peptides play a role in modulating the insulin-stimulated sympathetic nerve response.

Chronic nitric oxide deficiency is associated with altered leutinizing hormone and follicle-stimulating hormone release in ovariectomized rats.

Nitric oxide (NO) synthase (NOS) has been found in the gonadotrophs and folliculo-stellate cells of the anterior pituitary. Previous observations from our laboratory suggest that NO may play a role in regulating gonadotropin secretion. Because estrogen secretion by the ovary can influence gonadotropin secretion, we investigated the hypothesis that chronic in vivo NO deficiency has a direct estrogen-independent effect on luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion. Chronic NO deficiency was induced by adding an NOS inhibitor, N-nitro-L-arginine (L-NNA, 0.6 g/l) to the drinking water of ovariectomized (OVX) rats. The control OVX rats were untreated. After 6-8 weeks, the animals were sacrificed, and the pituitaries were removed and perfused continuously for 4 hr in the presence of pulsatile gonadotropin-releasing hormone (GnRH, 500 ng/pulse) every 30 min. S-Nitroso-L-acetyl penicillamine (SNAP, an NO donor, 0.1 mM) or L-nitro-arginine methyl ester (L-NAME, an NOS inhibitor, 0.1 mM) was added to the media and perfusate samples were collected at 10-min intervals. GnRH-stimulated LH and FSH levels were significantly lower in pituitaries from OVX/NO-deficient pituitaries compared with pituitaries from the OVX control group. The addition of SNAP significantly decreased LH and FSH secretion by pituitaries from OVX control animals, but significantly increased their secretion by pituitaries from the OVX/NO-deficient animals. L-NAME also suppressed LH and FSH secretion by pituitaries from the OVX control animals and stimulated their release by pituitaries from the NO-deficient/OVX animals. Immunohistochemistry of frontal sections through the hypothalamus demonstrated that OVX/NO deficiency is associated with increased GnRH in the median eminence. We conclude that NO has a chronic stimulatory effect on LH and FSH release and the subsequent altered secretory responsiveness to NO agonist or antagonist is the result of chronic NO suppression.

The effects of beta-endorphin (beta-END) on cardiovascular and behavioral dynamics in conscious rats.

Beta-endorphin (beta-END) a product of the proopiomelanocortin (POMC) has been demonstrated to play a role in the regulation of metabolic and autonomic responses. Recent studies have suggested the involvement of the endogenous opioid system in cardiovascular control. Previous studies conducted in our laboratory using anesthetized animals investigated the actions of beta-END and other POMC derived peptides on sympathetic and cardiovascular dynamics. In this study, we determined both the acute and chronic effects of beta-END on cardiovascular and behavioral dynamics in conscious unrestrained rats using radio-telemetry. Animals were instrumented with a radio-telemetry transmitter in the abdominal cavity and the attached catheter inserted into the femoral artery for recording of cardiovascular dynamics and activity. They were subsequently implanted with intracerebroventricular (ICV) cannulas. The acute ICV administration of beta-END significantly increased the mean arterial pressure (MAP) and heart rate (HR) compared to controls. The cardiovascular responses returned toward control levels after 2 h. In contrast, the chronic infusion of beta-END significantly decreased the MAP and HR during both the active and inactive phase. Chronic beta-END administration also decreased physical activity. Food intake was increased initially and later declined and water consumption followed a similar pattern. We conclude that in the conscious unrestrained animal the acute administration of beta-END increases MAP and HR while the chronic infusion of beta-END decreases MAP, HR, physical activity, and stimulate a short-term increase in food and water intake.

High fat feeding is associated with increased blood pressure, sympathetic nerve activity and hypothalamic mu opioid receptors.

Obesity and high fat diets are associated with an increased prevalence of diabetes, cardiovascular disease, and hypertension. However, the mechanism(s) linking obesity and high fat diet to these metabolic and cardiovascular disorders are not fully elucidated. Leptin stimulates the formation of pro-opiomelanocortin and its products. The stimulation of the central nervous system (CNS) opioids and their receptors is associated with an increase in cardiovascular dynamics. In this study we hypothesized that obesity changed the CNS opioids and their receptors that could play a role in altered cardiovascular and autonomic nervous regulation in obesity. Male Wistar rats were fed either a high fat (HF) or regular chow (control) diet. After 12 weeks, rats were anesthetized and instrumented to record mean arterial pressure (MAP) and renal sympathetic nerve activity (RSNA). A blood sample was collected and plasma glucose, insulin, leptin, beta-endorphins were measured. The brains were subsequently processed for immunohistochemistry and in situ hybridization. The HF rats were larger and had a greater percentage of body fat. Leptin and insulin levels were also higher in the HF animals. Basal MAP and RSNA were significantly higher in HF rats. Additionally, immunohistochemistry and in situ hybridization demonstrated that HF rats had increased hypothalamus mu opioid receptors compared to controls. These studies suggest that HF feeding is associated with increased body fat, plasma leptin, insulin, and hypothalamic mu opioid receptors. The increased mu opioid receptors may contribute to the higher MAP and RSNA observed in HF animals.

Diabetes enhances apoptosis induced by cerebral ischemia.

The aim of this study is to explore the mechanism by which diabetes exaggerates cerebral stroke and its outcome. Since ischemia can be related to not only necrosis but apoptosis as well, we compared the development of apoptosis in STZ-diabetic rats and STZ-diabetic rats subjected to occlusion of the middle cerebral artery (MCA). 24-48 hr following MCA occlusion the animals were killed, the brain removed and prepared for evaluation by several indexes of apoptosis: nucleosomal DNA fragmentation, TUNEL staining, activation of caspase-3 and alteration in the expression of Bax and Bcl2. DNA fragmentation was not detected in the cortex of normal and diabetic animals, but was evident following MCA occlusion in diabetic rats. Bax expression was increased in the cortex of normal rats following MCA occlusion and this expression was further increased in the cortex of MCA occluded diabetic rats. Bcl2 expression was not changed in any of the groups. In the hippocampus, DNA fragmentation was not evident in control rats but was observed in diabetic rats. Ischemic injury did not enhance DNA laddering in diabetic animals. The expression of Bax was increased in diabetic rats but was not increased following MCA occlusion. Bcl2 expression was not changed by ischemia in any of the animal models. These data suggest that diabetes may enhance the development of stroke via increased cortical apoptotic activity but this was not additive in the hippocampus following ischemic injury.

Expression of cAMP-responsive element modulator (CREM) in rat testes following chronic cocaine administration.

OBJECTIVE: c-AMP-responsive element modulator (CREM), one of the nuclear factors involved in the regulation of gene expression by cAMP, has an important role in spermatogenesis. Our recent study has shown that chronic administration of cocaine to male rats results in disruption of spermatogenesis, including reduction of germ cells. As a further step toward understanding this process, we have studied the role of CREM in cocaine-induced testicular damage. MATERIALS AND METHODS: Sprague-Dawley rats were administered cocaine hydrochloride subcutaneously daily for 90 days. Control animals received equal volumes of normal saline daily for 90 days. Testes were removed after 15, 30, 90 days of cocaine administration. Total RNA was extracted from the testes and subjected to RT-PCR. Testicular tissue was also homogenized in a lysis buffer, and Western blotting was performed using anti-CREM antibody. RESULTS: RT-PCR analysis detected a single fragment of approximately 520 base pairs (bp) in control testes at all time points. The cocaine-treated testes showed reduced expression of CREM fragment. Western blot analysis using CREM antibodies confirmed the RNA data. There were reduced CREM proteins in the cocaine-treated testes compared with controls. CONCLUSIONS: The CREM gene is essential for spermatogenesis. Our results indicate that the reduction in testicular CREM expression may be one of the mechanisms responsible for disruption or impairment of spermatogenesis in the testes following chronic cocaine administration.

Cigarette smoking induces apoptosis in rat testis.

In a previous studywe demonstrated the deleterious effect of cigarette smoke on spermatogenesis in the testis of peripubertal Sprague-Dawley rats. In this study we investigated the development of apoptosis as a possible contributing factor to the pathogenic mechanism underlying these effects. Peripubertal rats were exposed to cigarette smoke with the Walton Horizontal Smoking Machine. Similarly, age-matched control rats were exposed to room air with the smoking machine. Rats from both groups were sacrificed after 45 days of treatment and the testes were removed. Testes were stained utilizing the terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) staining technique. DNA fragmentation was further evaluated using gelectrophoresis. There was a significant increase in the incidence of apoptosis in the treated group compared to the control group as demonstrated by the larger amount of tubules containing > or = 3apoptotic bodies in the smoke-exposed group, that is, 36% versus 14% in the control group (p < 0.05). Agarose gel electrophoresis demonstrated the DNA ladder in the treated group but not in the control animals. In conclusion, chronic cigarette smoke induces apoptosis in the rat testis. Apoptosis may be one of the pathogenic mechanisms responsible for defective spermatogenesis in the rat following chronic cigarette smoking.

mRNA redistribution during permanent focal cerebral ischemia.

Translation arrest occurs in neurons following focal cerebral ischemia and is irreversible in penumbral neurons destined to die. Following global cerebral ischemia, mRNA is sequestered away from 40S ribosomal subunits as mRNA granules, precluding translation. Here, we investigated mRNA granule formation using fluorescence in situ histochemistry out to 8 h permanent focal cerebral ischemia using middle cerebral artery occlusion in Long Evans rats with and without diabetes. Neuronal mRNA granules colocalized with PABP, HuR, and NeuN, but not 40S or 60S ribosomal subunits, or organelle markers. The volume of brain with mRNA granule-containing neurons decreased exponentially with ischemia duration, and was zero after 8 h permanent focal cerebral ischemia or any duration of ischemia in diabetic rats. These results show that neuronal mRNA granule response has a limited range of insult intensity over which it is expressed. Identifying the limits of effective neuronal stress response to ischemia will be important for developing effective stroke therapies.

Insulin like growth factor-1 (IGF-1) decreases ischemia-reperfusion induced apoptosis and necrosis in diabetic rats.

Previous evidence supports the view that insulin, as well as insulin like growth factor-1 (IGF-1) provides neurotropic support for neurons in the central nervous system (CNS) and peripheral nervous system (PNS). In the present study we evaluated the effects of the intravenous infusion of IGF-1 on both necrosis and apoptosis in the CNS of streptozotocin induced diabetic animals before and/or following middle cerebral artery occlusion (MCAO) with reperfusion. The lesion volume was used as an index of necrosis and the sensorimotor cortex (layers 5 and 6) as well as the CA1 and CA3 regions of the hippocampus were analyzed for apoptosis using TUNEL staining and Caspase-3 immunoreactivity. A large lesion volume was produced in diabetic animals after 2-h MCAO and 24-h reperfusion. Diabetic animals also had an elevated basal level of apoptotic cells that are bilaterally distributed. Apoptosis was further increased over basal after 2-h MCAO and 24-h reperfusion. The acute administration of IGF-1 30-min before or 2 h after MCAO followed by 24-h reperfusion decreased the lesion volume as well as the number of apoptotic cells in the cortical penumbra. Apoptosis as assessed by TUNEL and caspase-3 immunoreactivity was decreased in select sensorimotor cortex and hippocampal areas. We conclude that treatment with IGF-1 before or after ischemic insult significantly decreases both lesion volume and apoptosis in selected areas of the cortex and hippocampus.

The effect of high fat-induced obesity on cardiovascular and physical activity and opioid responsiveness in conscious rats.

Both obesity and increased endorphin production are associated with an increase in blood pressure. We have previously demonstrated that the acute and chronic central nervous system (CNS) administration of beta-endorphin can increase or decrease blood pressure, respectively. Also high fat (HF) diet-induced obesity is associated with increased hypothalamic mu opioid receptors and increased blood pressure in response to ss-endorphins. In this study we investigated the effect of high fat diet-induced obesity on blood pressure, heart rate, and physical activity as well as determined the effect of mu opioids in unanesthetized rats. Male Wistar rats were implanted with a radiotelemetry transmitter to record cardiovascular dynamics and activity. They were fed either a HF diet (HF; 59% fat by caloric content, soy bean oil) or regular chow (control; 12% fat by caloric content). HF rats had higher body weights and their total caloric intake was greater than controls. The systolic blood pressures (SBP) were greater in the HF-obese rats. After 12-13 weeks the rats were infused chronically with a mu opioid agonist (D)-Ala(2), N-Me-Phe(4), Gly(5)-ol]-ENKEPHALIN (DAMGO) or a mu opioid antagonist ss-funaltrexamine (beta-FNA) via intracerebroventricular cannula. DAMGO increased the SBP and heart rate in controls, but not in HF obese rats. DAMGO did not affect physical activity; beta-FNA decreased SBP and increased HR in controls. We concluded that HF rats consumed more calories, gained more weight, and had higher SBP. However, the responsiveness to the mu-receptor agonist was not higher in the HF rats.

The insulin-mediated vascular and blood pressure responses are suppressed in CGRP-deficient normal and diabetic rats.

BACKGROUND: Calcitonin gene-related peptide (CGRP) is extensively localized in the perivascular or periadventitia nerves throughout the body. CGRP is a potent vasodilator and its release is associated with dilation of these blood vessels. The present study investigated the contribution of the CGRP-mediated vasodilation to the insulin-induced vasodilatory response. METHODS: Male Wistar rats were treated with capsaicin (50 mg/kg) at 1-3 days of age to ablate the CGRP-containing neurons. After 8 weeks some animals were made diabetic using streptozotocin. Vehicle-treated animals were used as controls. At 12-13 weeks the animals were fasted, anesthetized with chloralose/urethane and instrumented for recording of cardiovascular dynamics. RESULTS: Body weights and basal, insulin, glucose, mean arterial pressure (MAP), heart rate (HR), and vascular flows were not different in CGRP-deficient rats versus controls. Insulin infusion significantly decreased the MAP in vehicle-treated controls but this response was completely attenuated in CGRP-deficient rats. The decreased response to insulin was associated with a diminished vascular dilatory response in the iliac, renal, and superior mesenteric vessel beds. When insulin was infused in CGRP-deficient diabetic animals there was also a diminished response. Diabetes resulted in an increased renal vascular flow in response to insulin. CONCLUSIONS: From the present studies we conclude that the insulin-mediated vasodilation was due, in part, to the stimulation of perivascular nerves to release CGRP, and the action of CGRP on vascular smooth muscle enhanced directly or indirectly the vasodilatory response to insulin.

Cardiovascular action of insulin-like growth factor-1 is not mediated by calcitonin gene-related peptide neurons.

Calcitonin gene-related peptide (CGRP) is a potent vasodilator located in the peripheral nerves including the perivascular nerves. Previous studies in our laboratory determined that the vasodilatory action of insulin is mediated in part by CGRP-containing neurons. Since insulin-like growth factor-1 (IGF-1) and insulin share molecular and receptor structural similarity as well as functional similarity, we investigated the role of the CGRP-containing neurons in IGF-1-mediated vasodilation. Wistar rats were made CGRP deficient by treatment with capsaicin (50 mg/kg) 1-3 d after birth. Vehicle-treated controls and CGRP-deficient rats were maintained for 12 to 13 wk. At this time rats were fasted overnight, anesthetized with urethane and chloralose, and prepared for cardiovascular recordings. The basal mean arterial pressure (MAP) was higher in CGRP-deficient rats when compared with controls. The infusion of IGF-1 resulted in an equivalent decrease in MAP in both the CGRP-deficient and control rats. IGF-1 infusion did not change the heart rate in control rats but decreased it in CGRP-deficient rats. IGF-1 also increased flow as determined by conductance in the iliac, renal, and superior mesenteric vascular beds in both vehicle controls and CGRP-deficient rats. We concluded that unlike insulin the IGF-1-mediated vasodilatory response is not mediated by the CGRP-dependent perivascular neurons.

The effects of middle cerebral artery occlusion on central nervous system apoptotic events in normal and diabetic rats.

Apoptosis and neural degeneration are characteristics of cerebral ischemia and brain damage. Diabetes is associated with worsening of brain damage following ischemic events. In this study, the authors characterize the influence of focal cerebral ischemia, induced by middle cerebral artery occlusion, on 2 indexes of apoptosis, TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine 5-triphosphate nick end-labeling) staining and caspase-3 immunohistochemistry. Diabetes was induced in normal rats using streptozotocin and maintained for 5 to 6 weeks. The middle cerebral artery of both normal and diabetic rats was occluded and maintained from 24 or 48 hours. Sham-operated normal and diabetic animals served as controls. Following 24 to 48 hours of occlusion, the animals were sacrificed and the brains were removed, sectioned, and processed for TUNEL staining or caspase-3 immunohistochemistry. Middle cerebral artery occlusion in normal rats was associated with an increase in the number of both TUNEL-positive and caspase-3-positive cells in selected brain regions (hypothalamic preoptic area, piriform cortex, and parietal cortex) when compared to nonoccluded controls. Diabetic rats without occlusion showed significant increases in both TUNEL-positive and caspase-3-positive cells compared to normal controls. Middle cerebral artery occlusion in diabetic rats resulted in increases in TUNEL-positive as well as caspase-3-positive cells in selected regions, above those seen in nonoccluded diabetic rats. Both TUNEL staining and caspase-3 immunohistochemistry revealed that the number of apoptotic cells in diabetic animals tended to be greatest in the preoptic area and parietal cortex. The authors conclude that focal cerebral ischemia is associated with a significant increase in apoptosis in nondiabetic rats, and that diabetes alone or diabetes plus focal ischemia are associated with significant increases in apoptotic cells.