Cardiovascular effects of the intracerebroventricular injection of adrenomedullin: roles of the peripheral vasopressin and central cholinergic systems

Our objective was to investigate in conscious Sprague-Dawley (6-8 weeks, 250-300 g) female rats (N = 7 in each group) the effects of intracerebroventricularly (icv) injected adrenomedullin (ADM) on blood pressure and heart rate (HR), and to determine if ADM and calcitonin gene-related peptide (CGRP) receptors, peripheral V1 receptors or the central cholinergic system play roles in these cardiovascular effects. Blood pressure and HR were observed before and for 30 min following drug injections. The following results were obtained: 1) icv ADM (750 ng/10 µL) caused an increase in both blood pressure and HR (DMAP = 11.8 ± 2.3 mmHg and ΔHR = 39.7 ± 4.8 bpm). 2) Pretreatment with a CGRP receptor antagonist (CGRP8-37) and ADM receptor antagonist (ADM22-52) blocked the effect of central ADM on blood pressure and HR. 3) The nicotinic receptor antagonist mecamylamine (25 µg/10 µL, icv) and the muscarinic receptor antagonist atropine (5 µg/10 µL, icv) prevented the stimulating effect of ADM on blood pressure. The effect of ADM on HR was blocked only by atropine (5 µg/10 µL, icv). 4) The V1 receptor antagonist [β-mercapto-β-β-cyclopentamethylenepropionyl¹, O-me-Tyr²,Arg8]-vasopressin (V2255; 10 µg/kg), that was applied intravenously, prevented the effect of ADM on blood pressure and HR. This is the first study reporting the role of specific ADM and CGRP receptors, especially the role of nicotinic and muscarinic central cholinergic receptors and the role of peripheral V1 receptors in the increasing effects of icv ADM on blood pressure and HR.

Cardiovascular effects of the intracerebroventricular injection of adrenomedullin: roles of the peripheral vasopressin and central cholinergic systems.

Our objective was to investigate in conscious Sprague-Dawley (6-8 weeks, 250-300 g) female rats (N = 7 in each group) the effects of intracerebroventricularly (icv) injected adrenomedullin (ADM) on blood pressure and heart rate (HR), and to determine if ADM and calcitonin gene-related peptide (CGRP) receptors, peripheral V1 receptors or the central cholinergic system play roles in these cardiovascular effects. Blood pressure and HR were observed before and for 30 min following drug injections. The following results were obtained: 1) icv ADM (750 ng/10 µL) caused an increase in both blood pressure and HR (DMAP = 11.8 ± 2.3 mmHg and ΔHR = 39.7 ± 4.8 bpm). 2) Pretreatment with a CGRP receptor antagonist (CGRP8-37) and ADM receptor antagonist (ADM22-52) blocked the effect of central ADM on blood pressure and HR. 3) The nicotinic receptor antagonist mecamylamine (25 µg/10 µL, icv) and the muscarinic receptor antagonist atropine (5 µg/10 µL, icv) prevented the stimulating effect of ADM on blood pressure. The effect of ADM on HR was blocked only by atropine (5 µg/10 µL, icv). 4) The V1 receptor antagonist [ß-mercapto-ß-ß-cyclopentamethylenepropionyl¹, O-me-Tyr²,Arg8]-vasopressin (V2255; 10 µg/kg), that was applied intravenously, prevented the effect of ADM on blood pressure and HR. This is the first study reporting the role of specific ADM and CGRP receptors, especially the role of nicotinic and muscarinic central cholinergic receptors and the role of peripheral V1 receptors in the increasing effects of icv ADM on blood pressure and HR.

Evaluation of platelet function and lack of response to epinephrine in pregnant women.

Previous studies in healthy subjects have demonstrated a lack of response of platelets to epinephrine at a rate of 16-40% on an aggregometer. An association between the increased procoagulant factors during pregnancy and venous thromboembolism is known, and it has also been shown that prolactin levels increase platelet aggregation. We evaluated whether platelet functions in pregnant women and also assessed the lack of response to epinephrine during this period. We compared 27 healthy and volunteering pregnant women with 26 similar control subjects. Platelet functions were assessed with an aggregometer and a Platelet Function Analyzer (PFA-100). Less than 40% response to epinephrine on the aggregometer was defined as an impaired epinephrine response. The aggregation response of epinephrine was normal in 25 of the 27 pregnant women, while two of them showed a late-rising response. Eight of the 26 subject control group (30.8%) showed an impaired response to epinephrine. When we compared the 25 pregnant and 18 control subjects with normal aggregation responses, the maximum aggregation responses to ADP and epinephrine, and the Col/Epi and Col/ADP cartridge closure time values were significantly lower in pregnant women. There were no difference between second and third trimesters as regards platelet function parameters. The fact that no impaired response to epinephrine was detected in pregnant women while a 30% rate was observed in non-pregnant women indicates that the platelet malfunction caused by a disorder in the Gi protein and intracellular mechanisms is bypassed during pregnancy thanks to some physiological changes.

Propofol versus isoflurane anesthesia under hypothermic conditions: effects on intracranial pressure and local cerebral blood flow after diffuse traumatic brain injury in the rat.

BACKGROUND: The aim of this study was to compare the cerebral protective effects of two known protective anesthetics, isoflurane and propofol, when these were used in combination with moderate hypothermia (33-34 degrees C) after diffuse traumatic brain injury (TBI) in the rat. We assessed cerebral protection by measuring local cerebral blood flow (LCBF), mean arterial blood pressure (MABP), cerebral perfusion pressure (CPP) and intracranial pressure (ICP). METHODS: Sixteen female Wistar rats weighing 275 to 350 g were anesthetized and subjected to an accelerated-impact weight-drop model of diffuse TBI. Hypothermia (33-34 degrees C) was induced 45 minutes after TBI (baseline), and was maintained for 180 minutes. The isoflurane group (n = 8) received 70% N(2)O in O(2), and isoflurane at 0.9 +/- 0.04%. The propofol group (n = 8) received 70% N(2)O in O(2) and a propofol infusion (12 mg/kg/hr). LCBF was measured by laser Doppler flowmeter. MABP, ICP, and brain and rectal temperatures were measured every 15 minutes from baseline through 180 minutes. Blood gas and hematocrit testing was also done at baseline and every 60 minutes thereafter to assess the animals' physiological state. RESULTS: In the isoflurane group, MABP and CPP decreased significantly from baseline to 180 minutes (p < 0.05 and p < 0.01, respectively), and MABP was significantly lower than the pressure in the propofol group from 45 minutes through 180 minutes (p < 0.05, p < 0.01). ICP and LCBF remained unchanged in this group. In the propofol group, from baseline to 180 minutes, CPP increased to maximum 120 +/- 8 mmHg at 75 minutes from 98 +/- 5 mmHg (p < 0.05) and ICP fell from 18 +/- 2 mmHg to 7 +/- 1 mmHg (p < 0.01); and the latter was significantly lower than ICP in the isoflurane group (p < 0.05, p < 0.01, p < 0.001). LCBF in this group was significantly higher than LCBF in the isoflurane group in the last 30 minutes of the experiment (p < 0.05). The propofol group showed no change in MABP over the course of the experiment. CONCLUSION: In the clinical setting, propofol anesthesia may be better for use in combination with hypothermia in cases of traumatic brain injury, as it reduces ICP and increases CPP under these conditions.

A central link between angiotensinergic and cholinergic systems; role of vasopressin.

Participation of central cholinergic system in the effects of intracerebroventricular (i.c.v.) injection of angiotensin II (Ang II) on blood pressure and heart rate was studied in conscious, freely moving rats. Ang II dose-dependently increased blood pressure and decreased heart rate. Both atropine and mecamylamine (i.c.v.) pretreatments prevented the cardiovascular effects of Ang II. Pretreatment with a vasopressin V1 antagonist also prevented the cardiovascular responses to Ang II. Our data suggest that the central pressor effect of Ang II is mediated in part by central acetylcholine via both muscarinic and nicotinic receptors, and vasopressin participates in this effect through V1 receptors.

Central injection of captopril inhibits the blood pressure response to intracerebroventricular choline

In the present study, we investigated the involvement of the brain renin-angiotensin system in the effects of central cholinergic stimulation on blood pressure in conscious, freely moving normotensive rats. In the first step, we determined the effects of intracerebroventricular (icv) choline (50, 100 and 150 µg) on blood pressure. Choline increased blood pressure in a dose-dependent manner. In order to investigate the effects of brain renin-angiotensin system blockade on blood pressure increase induced by choline (150 µg, icv), an angiotensin-converting enzyme inhibitor, captopril (25 and 50 µg, icv), was administered 3 min before choline. Twenty-five µg captopril did not block the pressor effect of choline, while 50 µg captopril blocked it significantly. Our results suggest that the central renin-angiotensin system may participate in the increase in blood pressure induced by icv choline in normotensive rats.

Central injection of captopril inhibits the blood pressure response to intracerebroventricular choline.

In the present study, we investigated the involvement of the brain renin-angiotensin system in the effects of central cholinergic stimulation on blood pressure in conscious, freely moving normotensive rats. In the first step, we determined the effects of intracerebroventricular (icv) choline (50, 100 and 150 microg) on blood pressure. Choline increased blood pressure in a dose-dependent manner. In order to investigate the effects of brain renin-angiotensin system blockade on blood pressure increase induced by choline (150 microg, icv), an angiotensin-converting enzyme inhibitor, captopril (25 and 50 microg, icv), was administered 3 min before choline. Twenty-five microg captopril did not block the pressor effect of choline, while 50 microg captopril blocked it significantly. Our results suggest that the central renin-angiotensin system may participate in the increase in blood pressure induced by icv choline in normotensive rats.

Acute and delayed vasoconstriction after subarachnoid hemorrhage: local cerebral blood flow, histopathology, and morphology in the rat basilar artery.

The decreased local cerebral blood flow (LCBF) and cerebral ischemia that occur after subarachnoid hemorrhage (SAH) may be caused by acute and/or delayed vasospasm. In 36 Sprague-Dawley (350-450 g) rats SAH was induced by transclival puncture of the basilar artery. Mean arterial blood pressure (MABP), LCBF, intracranial pressure (ICP), and cerebral perfusion pressure (CPP) were measured in all rats for 30 min before and 60 min after SAH was induced. One set of control (n : 7) and experimental animals (n : 7) was sacrificed after the 60 min of initial post-hemorrhage measurements were recorded. Four days after SAH induction, LCBF and MABP were measured again for 60 min in subgroups of surviving experimental rats (n : 7) and control rats (n : 7). Histopathologic and morphologic examinations of the basilar artery were performed in each subgroup. There was a sharp drop in LCBF just after SAH was induced (55.50 +/- 11.46 mlLD/min/100 g and 16.1 +/- 3.6 mlLD/min/100 g for baseline and post-SAH, respectively; p < 0.001). The flow then gradually increased but had not returned to pre-SAH values by 60 min (p < 0.05). At 4 days after SAH induction, although LCBF was lower than that observed in the control group and pre-SAH values, it was not significantly different from either of these flow rates (p > 0.05). ICP (baseline 7.05 +/- 0.4 mmHg) increased acutely to 75.2 +/- 7.1 mmHg, but returned to normal levels by 60 min after SAH. CPP (baseline 84.5 +/- 6.3 mmHg) dropped accordingly (to 18.6 +/- 3.1 mmHg), and then increased, reaching 72.2 +/- 4.9 mmHg at 60 min after SAH (p > 0.05). Examinations of the arteries revealed decreased inner luminal diameter and distortion of the elastica layer in the early stage. LCBF in nonsurviver rats (n : 8) was lower than that in the animals that survived (p < 0.01). At 4 days post-hemorrhage, the rats' basilar arteries showed marked vasculopathy. The findings showed that acute SAH alters LCBF, ICP, and CPP, and that decreased LCBF affects mortality rate. Subsequent vasculopathy occurs in delayed fashion, and this was observed at 4 days after the hemorrhage event.

Effects of interrupted and uninterrupted occlusion of the basilar artery on cerebral blood flow, and on neurological and histological outcome in rats with subarachnoid hemorrhage.

Most neurosurgeons consider temporary vessel occlusion for aneurysmal clipping an effective technique that facilitates dissection between the aneurysm and the parent vessel. It is generally believed that repeated short periods of cerebral ischemia are safer for the brain than a single long episode. The aim of this study was to identify whether interrupted and uninterrupted vessel occlusion differs with regard to changes in brain tissue and cerebral hemodynamics after subarachnoid hemorrhage (SAH). Fifty Spraque Dawley rats (300-350 g) were placed under general anaesthesia and ventilated. The basilar artery was exposed through a transclival approach. Baseline local cerebral blood flow (LCBF) values was measured, and then the basilar artery was punctured, causing subarachnoid hemorrhage (SAH). Group I (n = 24) was subjected to 60 min of interrupted basilar artery occlusion, defined as 5 min of reperfusion after every 10 min of occlusion, group II (n = 26) 60 min of uninterrupted artery occlusion. Three days after completion of the experiment, each rat was neurologically evaluated and decapitated. Coronal brain slices were obtained and stained to assess infarct volume. Immediately after SAH, LCBF fell by 58% in group I, and by 52% in group II. In group I, each ischemic insult brought a similar reduction in LCBF, and after each release of the occlusion there was a rapid rise in flow. In group II, the LCBF values dropped initially and remained at low levels until the end of the study. The 2,3,5 triphenyltetrazolium chloride stained sections showed similar volumes of brainstem infarction in both groups (38.3 +/- 9.2 mm3 vs. 34.3 +/- 8.7 mm3, respectively; p > 0.05). The results suggest that there is no neuroprotective advantage to either interrupted or uninterrupted temporary blockage of blood flow during neurovascular procedures after SAH in the basilar artery region.

Neuroprotective effects of MK 801 and hypothermia used alone and in combination in hypoxic-ischemic brain injury in neonatal rats.

Although accumulating evidence suggests that increased extracellular glutamate concentrations may play an important role in hypoxic-ischemic brain injury, dopamine and other catecholamines also seem to be involved. The N-methyl-D-aspartate receptor antagonist MK 801 and moderate hypothermia (32-34 degrees C) are each known to be neuroprotective, but their combined effect on the release and metabolism of neurotransmitters is unknown. Seven-day-old pups (n: 150) underwent right common carotid artery ligation to induce hemispheric ischemia, and were later subjected to 120 minutes of hypoxia with 8% O2 and 92% N2O. Half the rats (Group I, n: 74) were subjected to normothermic conditions throughout the hypoxic period. Moderate hypothermia (30-32 degrees C) was induced in the other pups (Group II, n: 76) immediately after artery occlusion, and was maintained throughout the hypoxic period. Prior to inducing hypoxia, half of the rats in each group (Groups IA and IIA) received vehicle solution (0.9% NaCI) and the other rats (Groups IB and IIB) received MK 801 (0.5 mg/kg) subcutaneously at 45 and 120 minutes after occlusion. Intracerebral temperature was recorded every 15 minutes after occlusion. Infarct area (n: 40) was calculated after staining with 2% 2,3,5 triphenyltetrazolium chloride. Neuronal damage (n: 42) was assessed by quantifying CA1-CA3 neuronal loss at five hippocampal levels. The amount of damage to the monoamine system of the corpus striatum was determined based on the dopamine and 3,4 dihydroxyphenylacetic acid levels in the corpus striatum in both hemispheres (n: 46), as measured by high-pressure liquid chromatography and compared with normal control pups' values (n: 10). The normothermia/saline-treated pups had significantly larger infarct areas than the MK 801 only, hypothermia only, or MK 801/hypothermia combination groups. Neuropathological examination and striatal tissue monoamine data also confirmed marked neuronal damage in this group. Although MK 801 treatment alone resulted in significantly smaller infarct area and less tissue damage than was observed in the normothermia/saline-treated group, the moderate hypothermia and the MK 801/hypothermia combination treatment groups both exhibited better neuronal protection, especially in the corpus striatum. The rats that received combined treatment also had a significantly lower mortality rate.

Ischemic brain injury caused by interrupted versus uninterrupted occlusion in hypotensive rats with subarachnoid hemorrhage: neuroprotective effects of citicoline.

This study investigated the neuroprotection provided by cytidine 5'-diphosphocholine (citicoline) during interrupted and uninterrupted occlusion of the basilar artery after subarachnoid hemorrhage (SAH) in 121 hypotensive rats. Animals were anesthetized and the basilar artery was exposed through a transclival approach. Baseline local cerebral blood flow (LCBF) values were recorded, and then the basilar artery was punctured, causing SAH. Blood was drawn to induce hypotension [60-70 mmHg mean arterial blood pressure (MABP)]. Control rats received intraperitoneal (i.p.) injections of 0.5 ml saline immediately after SAH before hypotension induction and after 60 min of occlusion. Experimental rats received 400-mg/kg citicoline i.p. at the same time points. Control group I and treatment group III were subjected to 60 min of interrupted occlusion (5 min of reperfusion after each 10 min of occlusion). Control group II and treatment group IV were subjected to 60 min of uninterrupted occlusion. MABP and LCBF were recorded every 5 minutes. Brain edema was evaluated in seven rats from each group at 24 hours after ischemic injury. At 3 days after occlusion, another set of 28 rats was killed and coronal brain slices were stained to assess infarct volume. The groups' physiological and edema findings were similar. In all groups, LCBF fell immediately after SAH and remained below baseline throughout the experiment. In the citicoline-treated rats, arterial pressure increased significantly after 30-40 min of occlusion, and brain slices showed significantly smaller infarct volumes compared to control slices (p < 0.05). Mortality was significantly lower in the citicoline-treated animals (p < 0.001). The results suggest that citicoline provides significant neuroprotection during cerebral ischemia, and that it significantly reduces mortality. Part of the neuroprotective effect may be mediated by recovery of arterial pressure.

An anatomical and pathological evaluation of middle cerebral artery occlusion in rats.

Adult male Sprague-Dawley rats (n = 87) weighing 350-400 g were used for studying the anatomy of the horizontal segment of middle cerebral artery and infarct area after occlusion of the artery. In the experimental group (n = 27) middle cerebral artery was coagulated 3-4 mm length from the origin of the lateral striate arteries to the inferior cerebral vein and divided. Control rats (n = 20) had all the surgical procedures except occlusion. Another group of rats (n = 40) were used to determine the anatomical variations of middle cerebral artery after intracarotid carbon black injection. Five major patterns of middle cerebral artery were observed and two of them were major and constituted 92.5% of rats. Twenty-four hours after middle cerebral artery occlusion, all animals were neurologically evaluated. On the third day after occlusion the brains were stained with 2%, 2,3,5-triphenyltetrozolium chloride. The area of infarction was assessed by computerized analysis method. In our study after determining the variations of the middle cerebral artery and its branches in our strain of rats, we were able to achieve 92.5% grade III and IV infarcted area.

Role of intracerebroventricular vasopressin in the development of stress-induced gastric lesions in rats.

We investigated whether arginine vasopressin (AVP) has a central influence on the development of gastric mucosal lesions evoked by restraint-cold stress in rats. AVP and vasopressin V1 receptor antagonist were injected intracerebroventricularly (i.c.v.) and the rats were exposed to restraint-cold stress for five hours. After decapitation the stomachs were examined for gastric mucosal lesions which were evaluated according to an ulcer score. Three different doses of AVP and V1 receptor antagonist were administered in order to investigate the effects of exogenous and endogenous AVP on stress-induced gastric mucosal lesions. The intensity of gastric mucosal lesions was reduced when exogenous AVP was injected intracerebroventricularly. On the other hand, vasopressin V1 receptor antagonist, which was administered by the same route, augmented gastric mucosal lesions. Our findings indicate that AVP, injected centrally, plays a role in preventing the formation of gastric mucosal lesions induced by stress via a central V1 receptor.

The effect of endothelin-1 on vena jugularis thrombus model in rabbits.

Endothelins (ET) are the most important vasoconstrictors known, and administration results in contraction of vascular strips in man and experimental animals in vitro. We examined the effects of ET-1 on thrombus formation in rabbits. We used vasoconstrictor and thrombus forming agents and we selected an animal model, the vena jugularis thrombus model. In addition, intravascular endothelium was examined ultrastructurally. The ET-1 level is known to be high in patients with hypertension; if these patients also have atherosclerosis, then intravascular thrombus formation may increase. In the vena jugularis thrombus model, thromboplastin and ET-1 act synergistically to increase intravascular thrombus formation. On injection of ET-1 dose dependent vasoconstriction was shown in the vessel wall. Although similar maximal contraction is achieved, a decrease in vessel diameter is associated with increased potency of ET-1 and thromboplastin. The results suggest that ET-1 may regulate vascular tone through constriction of vessels.