Time-course study on doxorubicin-induced nephropathy and cardiomyopathy in male and female LOU/M/Wsl rats: lack of evidence for a causal relationship.

In a previous study on doxorubicin-induced cardiotoxicity in LOU/M/Wsl rats, severe nephropathy has been observed; therefore, the question was raised whether nephropathy adds to or even might be responsible for doxorubicin-induced cardiomyopathy in rats. For elucidation of this question, the temporal relationship between the onset of doxorubicin-induced cardiomyopathy and nephropathy was studied. In addition, examination was made of whether modifications of the treatment schedule could circumvent nephrotoxicity. Because preliminary studies had shown that female LOU/M/Wsl rats developed less doxorubicin-induced albuminuria, both male and female LOU/M/Wsl rats were treated with an iv dose of 1 mg doxorubicin/kg (body wt)/rat on five consecutive days and then weekly. Saline-treated animals served as controls. Albuminuria, serum albumin, and serum creatine levels were assessed weekly. For histologic examination, 5 male and 5 female rats were killed weekly. At day 14 and thereafter, doxorubicin-treated male rats showed albuminuria greater than or equal to 10 g/liter. Albuminuria of greater than or equal to 10 g/liter was not avoided by modifications of the treatment schedule. Female rats had on day 14 a urinary albumin level of 1.0-3.0 g/liter, yet reaching the level of greater than or equal to 10 g/liter at day 49. In male rats serum albumin levels decreased to levels below 10 g/liter (p less than .001 vs. finding for day 0); in contrast female rats maintained constant serum albumin levels till day 49. Serum creatine levels showed a tendency to rise, the values of male rats not being measured after day 28 due to hyperlipidemia; the levels of female rats increased from 37.8 +/- 3.0 mumol/liter to 53.7 +/- 2.5 mumol/liter on day 49 (P less than .001). At day 10 in male and female rats a grade 1-1.5 cardiomyopathy score, assessed according to the modified Billingham scoring system, was found, gradually increasing to grade 2.5-3 cardiomyopathy, both in males and females, on day 49. In male LOU/M rats the nephropathy developed steadily from day 14 and thereafter, whereas in females the rate of development of kidney damage was slower and at the end point of the study the severity of kidney lesions was less in comparison to that of the males. The onset of cardiomyopathy and nephropathy was simultaneous. It was concluded that cardiomyopathy observed in LOU/M rats is a phenomenon independent of nephropathy.

Reduced cardiotoxicity and nephrotoxicity with preservation of antitumor activity of doxorubicin entrapped in stable liposomes in the LOU/M Wsl rat.

LOU M/Wsl rats inoculated s.c. with 10(4) immunoglobulin immunocytoma cells have a palpable tumor at Day 17. Doxorubicin (DXR) has been entrapped in negatively charged liposomes (lip- DXR) composed of egg phosphatidylcholine, cholesterol, and phosphatidylserine and in positively charged liposomes (lip+ DXR) composed of phosphatidylcholine, cholesterol, and stearylamine. DXR, lip- DXR, and lip+ DXR were administered i.v. (0, 0.25, 0.5, 1.0, and 2.0 mg/kg) at Day 17 for 5 consecutive days and then weekly. Control animals showed progressive tumor growth leading to death 27 days after inoculation. Antitumor activity for all three preparations was dose dependent. DXR and lip- DXR showed the same antitumor activity; lip+ DXR had less antitumor activity. The overall survival of tumor-bearing animals treated with 2.0-mg/kg lip- DXR was significantly prolonged (p less than 0.05) in comparison to the animals treated with 2.0-mg/kg free DXR. Grade III cardiomyopathy was observed 47 days after treatment with free DXR; treatment with lip- DXR resulted in Grade I cardiomyopathy. In animals treated with 1.0-mg/kg and 2.0-mg/kg free DXR urinary albumin concentrations of 10 g/liter were observed. Treatment with 1.0-mg/kg lip- DXR and 1.0-mg/kg lip+ DXR resulted in urinary albumin concentration of less than 3.0 and less than 1.0 g/liter, respectively. Free DXR, 1.0 mg/kg, resulted in a decline of serum albumin concentration from 27.8 +/- 3.2 g/liter to 9.6 +/- 4.2 g/liter. No such decline was observed after treatment with lip- DXR and lip+ DXR. Treatment with a 1.0-mg/kg dose of free DXR resulted in severe glomerular and tubular lesions which were not found after treatment with 1.0-mg/kg lip- DXR and 1.0 mg/kg lip+ DXR. Administration of lip- DXR resulted in lower DXR levels in cardiac and renal tissue compared to administration of free DXR. After administration of lip+ DXR, very low tissue and tumor DXR levels were found. In conclusion, treatment with lip- DXR or lip+ DXR resulted in a prolonged survival, less albuminuria, and higher serum albumin levels. Also, fewer lesions in heart and kidney were found, correlating with lower DXR levels in these organs. Only lip- DXR had the same antitumor effect as free DXR.

Effect of gamma-melanocyte-stimulating hormones on baroreflex sensitivity and cerebral blood flow autoregulation in rats.

OBJECTIVE: In the present paper, we are interested in the effects of gamma-melanocyte-stimulating hormones (gamma-MSHs) on cardiovascular regulatory systems. METHODS: Mean arterial pressure (MAP), cerebral blood flow (CBF) and heart rate (HR) were measured in urethane-anaesthetised rats after intravenous administration of lysgamma(2)-MSH, gamma(2)-MSH, gamma(2)-MSH(6-12) or phenylephrine. RESULTS: The gamma-MSHs caused an increase in MAP, CBF and HR, whereas phenylephrine caused an increase in MAP and CBF and baroreceptor reflex-mediated bradycardia. All tested gamma-MSHs showed a significant impairment of the baroreceptor reflex sensitivity and CBF autoregulation as compared to the phenylephrine group. gamma(2)-MSH shows identical effects on the baroreceptor reflex and CBF as the endogenous occurring lysgamma(2)-MSH. In addition, the C-terminal fragment of gamma(2)-MSH, gamma(2)-MSH(6-12), induced similar effects as gamma(2)-MSH. The level of increase in MAP was comparable between the gamma-MSHs and the phenylephrine group. CONCLUSIONS: The present study suggests that gamma(2)-MSH and the shorter fragment gamma(2)-MSH(6-12) impair baroreceptor reflex sensitivity, due to a strong increase in sympathetic tone and/or change in baroreceptor reflex setpoint, and induce cerebrovasodilatation, which can counteract an autoregulation-mediated cerebrovasoconstriction due to systemic pressor effects. Furthermore, the results indicate that the C-terminal site of gamma(2)-MSH is relevant for its central-mediated inhibitory effects on the baroreceptor reflex and CBF.

In vivo cardiovascular reactivity and baroreflex activity in diabetic rats.

OBJECTIVES: Abnormalities of the cardiovascular system, e.g. impaired vasoreactivity and changes in baroreflex control of heart rate, are known to occur in experimental diabetes. It is not clear whether these cardiovascular dysfunctions are direct consequences of cardiovascular deficits and/or have autonomic neuropathy as a cause. METHODS: To differentiate between cardiovascular deficits or neuronal impairment as a cause for these cardiovascular dysfunctions, we tested the effects of the ACTH4-9 analogue, Org 2766, a neurotrophic compound without cardiovascular effects, on arterial pressure, heart rate and baroreflex control of heart rate. At 15 weeks, rats were made diabetic by injection of streptozotocin, and from 0-6, 6-12 or 12-18 weeks thereafter 3 groups of rats were treated with Org 2766. These effects were evaluated during phenylephrine-induced increases, and sodium nitroprusside-induced decreases, in blood pressure, in rats that had been diabetic for various periods (2-42 weeks). RESULTS: Throughout, both depressor response and maximal vasodilator activity in response to sodium nitroprusside were significantly (P < 0.05) reduced as compared to those of the non-diabetic controls. The pressor response of the diabetic rats to phenylephrine was only significantly (P < 0.05) reduced at 4, 6 and 12 weeks, and at 18 weeks, the diabetic rats were either hypo- or normoresponsive; Org 2766 did not restore the disturbed pressor response. From weeks 4 to 42 both maximal decrease in heart rate and sensitivity of baroreflex-mediated bradycardia in the diabetic rats were significantly less (P < 0.05) than those in the non-diabetic controls. Org 2766 restored the diminished baroreflex-mediated bradycardia of diabetic rats to non-diabetic control levels at 6 weeks, had an ameliorating effect at 12 weeks and no effect at 18 weeks. CONCLUSIONS: Time-dependent decreases in baroreflex sensitivity in diabetic rats was demonstrated and a much less steep decline of baroreflex sensitivity occurred in non-diabetic control rats. The ACTH4-9 analogue, Org 2766, when given immediately upon the induction of diabetes seem to delay the development of autonomic neuropathy, which suggests that cardiovascular factors appear to be of minor importance.

Middle cerebral artery occlusion in Wistar and Fischer-344 rats: functional and morphological assessment of the model.

Cerebral infarction volume after occlusion of a short proximal segment of the middle cerebral artery (MCA) is reported to be different in Wistar compared to Fischer-344 (F344) rats, in both size and variability. Knowledge about the cause of these differences might enable us to explain and perhaps reduce the variation in infarct volume and create a reproducible model of focal cerebral ischemia in the rat. We investigated in Wistar and F344 rats both the effect of occlusion of a long proximal MCA segment on cerebral infarction volume, visualized by magnetic resonance imaging and histology, and the morphology of the major cerebral arteries. Occlusion of a long proximal MCA segment resulted in a striatal and a small cortical infarction in Wistar and a striatal and sizable cortical infarction in F344 rats (as is the case after occlusion of a short proximal MCA segment). In Wistar rats, however, occlusion of a long proximal MCA segment strongly reduced the variability in infarction volume in comparison to occlusion of a small proximal MCA segment. Analysis of the morphology of the major cerebral arteries showed a significantly higher number of proximal side branches of the long proximal MCA segment in Wistar rates than in F344 rats. We conclude that after short-segment proximal MCA occlusion, extreme variability in cerebral infarction volume in Wistar rats compared to F344 rats may be attributable to a significantly greater number of proximal MCA side branches in Wistar rats than in F344 rats.

Relevance of the C-terminal Arg-Phe sequence in gamma(2)-melanocyte-stimulating hormone (gamma(2)-MSH) for inducing cardiovascular effects in conscious rats.

1. The cardiovascular effects by gamma(2)-melanocyte-stimulating hormone (gamma(2)-MSH) are probably not due to any of the well-known melanocortin subtype receptors. We hypothesize that the receptor for Phe-Met-Arg-Phe-amide (FMRFa) or Phe-Leu-Phe-Gln-Pro-Gln-Arg-Phe-amide (neuropeptide FF; NPFFa), other Arg-Phe containing peptides, is the candidate receptor. Therefore, we studied various Arg-Phe containing peptides to compare their haemodynamic profile with that of gamma(2)-MSH(6 - 12), the most potent fragment of gamma(2)-MSH. 2. Mean arterial pressure (MAP) and heart rate (HR) changes were measured in conscious rats after intravenous administration of gamma(2)-MSH related peptides. 3. Phe-Arg-Trp-Asp-Arg-Phe-Gly (gamma(2)-MSH(6 - 12)), FMRFa, NPFFa, Met-enkephalin-Arg-Phe-amide (MERFa), Arg-Phe-amide (RFa), acetyl-Phe-norLeu-Arg-Phe-amide (acFnLRFa) and desamino-Tyr-Phe-norLeu-Arg-Phe-amide (daYFnLRFa) caused a dose-dependent increase in MAP and HR. gamma(2)-MSH(6 - 12) showed the most potent cardiovascular effects (ED(50)=12 nmol kg(-1) for delta MAP; 7 nmol kg(-1) for delta HR), as compared to the other Arg-Phe containing peptides (ED(50)=177 - 292 nmol kg(-1) for delta MAP; 130 - 260 nmol kg(-1) for delta HR). 4. Peptides, which lack the C-terminal Arg-Phe sequence (Lys-Tyr-Val-Met-Gly-His-Phe-Arg-Trp-Asp-Arg-Pro-Gly (gamma(2)-pro(11)-MSH), desamino-Tyr-Phe-norLeu-Arg-[L-1,2,3,4 tetrahydroisoquinoline-3-carboxylic acid]-amide (daYFnLR[TIC]a) and Met-enkephalin (ME)), were devoid of cardiovascular actions. 5. The results indicate that the baroreceptor reflex-mediated reduction of tonic sympathetic activity due to pressor effects is inhibited by gamma(2)-MSH(6 - 12) and that its cardiovascular effects are dependent on the presence of a C-terminal Arg-Phe sequence. 6. It is suggested that the FMRFa/NPFFa receptor is the likely candidate receptor, involved in these cardiovascular effects.

Different cardiovascular profiles of three melanocortins in conscious rats; evidence for antagonism between gamma 2-MSH and ACTH-(1-24).

1. We investigated the effects of [Nle4,D-Phe7]alpha-melanocyte-stimulating hormone (NDP-MSH), adrenocorticotropin-(1-24) (ACTH-(1-24)) and gamma 2-MSH, three melanocortins with different agonist selectivity for the five cloned melanocortin receptors, on blood pressure and heart rate in conscious, freely moving rats following intravenous administration. 2. As was previously found by other investigators as well as by us gamma 2-MSH, a peptide suggested to be an agonist with selectivity for the melanocortin MC3 receptor, caused a dose-dependent, short lasting pressor response in combination with a tachycardia. Despite the fact that NDP-MSH is a potent agonist of various melanocortin receptor subtypes, among which the melanocortin MC1 receptor, it did not affect blood pressure or heart rate, when administered i.v. in doses of up to 1000 nmol kg-1. 3. ACTH-(1-24) caused a dose-dependent decrease in blood pressure in combination with a dose-dependent increase in heart rate in a dose-range from 15 to 500 nmol kg-1. The cardiovascular effects of ACTH-(1-24) were independent of the presence of the adrenals. 4. Pretreatment with ACTH-(1-24) caused a pronounced, dose-dependent parallel shift to the right of the dose-response curve for the pressor and tachycardiac effects of gamma 2-MSH. The antagonistic effect of ACTH-(1-24) was already apparent following a dose of this peptide as low as 10 nmol kg-1, which when given alone had no intrinsic hypotensive activity. 5. These results form further support for the notion that it is not via activation of one of the as yet cloned melanocortin receptors that gamma-MSH-like peptides increase blood pressure and heart rate. The cardiovascular effects of ACTH-(1-24) seem not to be mediated by the adrenal melanocortin MC3 receptors, for which ACTH-(1-24) is a selective agonist, or by adrenal catecholamines. 6. There appears to be a functional antagonism between ACTH-(1-24) and gamma 2-MSH, two melanocortins derived from a common precursor, with respect to their effect on blood pressure and heart rate. Whether this antagonism plays a (patho)physiological role remains to be shown.

Etomidate-anaesthesia, with and without fentanyl, compared with urethane-anaesthesia in the rat.

In spontaneously breathing rats, continuous infusion of etomidate with and without fentanyl caused a slight decrease in blood pressure and heart rate. Coadministration of fentanyl and etomidate in order to obtain full anaesthesia and analgesia resulted in respiratory depression. In artificially ventilated rats both etomidate as well as the anaesthetic combination caused a strong reduction in aortic flow and an increase in total peripheral resistance. A single infusion of etomidate did not change blood pressure. Etomidate combined with fentanyl reduced blood pressure. Under adjusted ventilation blood pressure, aortic flow, max(dF/dt) and heart rate were progressively reduced during a 4 h period. In contrast, urethane anaesthesia reduced aortic flow to a minor extent. Total peripheral resistance and max(dF/dt) were hardly affected. The slightly reduced blood pressure and blood gas variables remained stable during the experiment. From pharmacokinetic studies it was established that effective etomidate plasma levels were maintained constant during the experimental period. Pharmacokinetic interaction between etomidate and fentanyl did not occur. It is concluded that for anaesthesia of longer duration during cardiovascular experiments in rats, urethane is preferable to etomidate/fentanyl because it does not cause serious changes in basal haemodynamic variables.

Beneficial effect of the Ca2+ antagonist, nimodipine, on existing diabetic neuropathy in the BB/Wor rat.

1. Neuropathy is a frequently diagnosed complication of diabetes mellitus. Effective pharmacotherapy is not available. 2. The spontaneously diabetic BB/Wor rats develop secondary complications like neuropathy as do human diabetic patients. 3. BB/Wor rats treated with insulin via a subcutaneous implant show a significant impairment of sensory and motor nerve conduction velocity 6 weeks after the onset of diabetes mellitus. 4. Intraperitoneal treatment of diabetic BB/Wor rats with the Ca2+ antagonist, nimodipine (20 mg kg-1), from week 6 onwards every 48 h for a period of 6 weeks resulted in a significant increase of sensory and motor nerve conduction velocity. 5. Twelve weeks after the onset of diabetes mellitus BB/Wor rats show a 40% impairment of sciatic nerve blood flow as compared to the non-diabetic age-matched controls. Treatment with nimodipine (20 mg kg-1) from week 6 onwards significantly increased the sciatic nerve blood flow as compared to placebo-treated diabetic BB/Wor rats. 6. The adrenergic responsiveness of the vasa nervorum of the sciatic nerve to tyramine and phenylephrine was investigated as a parameter for autonomic neuropathy. 7. The fact that nimodipine treatment restored the reduced response to tyramine independently of the reduced postsynaptic phenylephrine responsiveness indicates that nimodipine improves adrenergic responsiveness mainly at the presynaptic level.

Mepyramine but not cimetidine or clobenpropit blocks pertussis toxin-induced histamine sensitization in rats.

The effects of pertussis toxin (PT) and the role of histaminergic H(1), H(2) and H(3) receptor blockade on the actions of histamine on blood pressure, heart rate, blood gas values, and mortality were studied in anaesthetized rats. Four days after treatment with PT, histamine dose-dependently decreased mean arterial blood pressure (MAP) and PT enhanced the histamine-induced decrease in MAP. In the PT but not in the inactivated PT (IPT) or saline treated group three out of six animals died after the highest dose of histamine (300 mg kg(-1), i.v.) In order to determine the type of histamine receptor that mediates HS, 4 days after PT the selective antagonists mepyramine (H(1)), cimetidine (H(2)) and clobenpropit (H(3)) were administered 20 min before the challenge with histamine. Mepyramine completely inhibited both the enhanced histamine-induced decrease in MAP and mortality brought about by PT. Cimetidine and clobenpropit had no protective effects, but rather enhanced the histamine-induced mortality elicited by PT. The present study shows that PT caused HS in rats which is primarily mediated via H(1) and secondarily via H(2) and H(3) receptors. These results are considered to be a first step in the elucidation of the mechanism(s) of the HS test used in the quality control of acellular pertussis vaccine.

ACE inhibitor-induced angioedema. Incidence, prevention and management.

Available information from 1980 to 1997 on angiotensin converting enzyme (ACE) inhibitor-induced angioedema and its underlying mechanisms are summarised and discussed. The incidence of angioedema is low (0.1 to 0.2%) but can be considered as a potentially life-threatening adverse effect of ACE inhibitor therapy. This adverse effect of ACE inhibitors, irrespective of the chemical structure, can occur early in treatment as well as after prolonged exposure for up to several years. The estimate incidence is quite underestimated. The actual incidence can be far higher because of poorly recognised presentation of angioedema as a consequence of its late onset in combination with usually long term therapy. Also, a spontaneous reporting bias can contribute to an actual higher incidence of this phenomenon. The incidence can be even higher (up to 3-fold) in certain risk groups, for instance Black Americans. Treatment includes immediate withdrawal of the ACE inhibitor and acute symptomatic supportive therapy followed by immediate (and long term) alternative therapy with other classes of drugs to manage hypertension and/or heart failure. Preclinical and clinical studies for the elucidation of the underlying mechanism(s) of ACE inhibitor-associated angioedema have not generated definite conclusions. It is suggested that immunological processes and several mediator systems (bradykinin, histamine, substance P and prostaglandins) are involved in the pathogenesis of angioedema. A great part of all reviewed reports suggest a relationship between ACE inhibitor-induced angioedema and increased levels of (tissue) bradykinin. However, no conclusive evidence of the role of bradykinin in angioedema has been found and an exclusive role of bradykinin seems unlikely. So far, no clear-cut evidence for an immune-mediated pathogenesis has been found. In addition, ACE gene polymorphism and some enzyme deficiencies are proposed to be involved in ACE inhibitor-induced angioedema. Progress in pharmacogenetic and molecular biological research should throw more light on a possible genetic component in the pathogenesis of ACE inhibitor-associated angioedema.

Synchronism of pressor response and grooming behavior in freely moving, conscious rats following intracerebroventricular administration of ACTH/MSH-like peptides.

After the i.c.v. administration of 300 pmol ACTH-(1-24) or [Nle4,D-Phe7]alpha-MSH, a long-lasting increase in blood pressure was observed synchronously with the incidence of excessive grooming. Two structurally related peptides with no grooming behavior-inducing potency, ACTH-(7-16)-NH2 and gamma 2-MSH, in doses of 300 and 500 pmol, respectively, caused a slight and short-lasting increase in blood pressure or had no effect, respectively. When the grooming behavior-inducing effect of ACTH-(1-24) was abolished, either by the prior manipulation of central dopaminergic neurotransmission by the i.c.v. administration of the dopamine receptor antagonist, haloperidol, or, due to the occurrence of single-dose tolerance to ACTH-(1-24), the pressor response was abolished as well. These data are in support of the postulate that the incidence of grooming behavior and the elevation of blood pressure are temporally associated and indicate that the two phenomena are causally related.

Differences in striatal extracellular amino acid concentrations between Wistar and Fischer 344 rats after middle cerebral artery occlusion.

We hypothesized that the interstrain difference between Wistar and Fischer-344 (F344) rats in cerebral infarction volume after proximal middle cerebral artery (MCA) occlusion might be explained by differences in excitotoxicity between both rat strains. Using microdialysis we measured during a 5 h period after MCA occlusion the release of aspartate, glutamate and taurine in the cerebral cortex and the striatum. The volume of striatal infarction was comparable in Wistar and F344 rats. We found, however, in Wistar rats a significantly higher striatal efflux of aspartate and glutamate than in F344 rats, whereas the striatal taurine efflux was of a similar magnitude in the two strains. Because of the (variably) smaller volume of cortical infarction in Wistar rats (than that in F344), the location of the microdialysis probe-membrane with respect to the area of cortical infarction differed between Wistar rats. Hence, a reliable comparison between the quantitative amount of amino acids in the dialysate from the cortical probes of both rat strains could not be made. These results, demonstrating differences in striatal excitotoxicity between Wistar and F344 rats after MCA occlusion, are the first to show interstrain differences in striatal pathophysiology of focal ischemia between these normotensive rat strains. They do however not explain why MCA occlusion results in a significantly different volume of cortical infarction between Wistar and F344 rats. The F344 strain will probably show in a more sensitive way, as compared to Wistar rats, neuroprotective effects of agents that diminish excitotoxic damage during focal cerebral ischemia.

Rat middle cerebral artery occlusion by an intraluminal thread compromises collateral blood flow.

We compared in Wistar rats collateral blood flow through leptomeningeal anastomoses after middle cerebral artery occlusion using craniotomy ('extravasal occlusion'), which results in a small volume of cerebral infarction, and after intraluminal thread occlusion ('intravasal occlusion'), which produces a large volume of cerebral infarction. Simultaneous laser-Doppler flowmetry with two probes placed on the cerebral cortex was used to measure and compare collateral blood flow. Extravasal occlusion caused a cortical blood flow reduction along a gradient in lateral direction, whereas blood flow reduction after intravasal occlusion was more uniformly distributed. It is concluded that permanent intravasal occlusion compromises collateral blood flow and therefore may not be a suitable model for measuring the ability of pharmacotherapeutic agents, if any, to improve collateral blood flow acutely after middle cerebral artery occlusion. The two models can be useful for testing drugs on parenchymal neuroprotective properties. Thereby, the intraluminal technique is preferred because of the possibility to study reperfusion damage when transient occlusion is applied.

Collateral hemodynamics after middle cerebral artery occlusion in Wistar and Fischer-344 rats.

We investigated whether the difference in infarction volume after occlusion of a long proximal segment of the middle cerebral artery between Wistar and Fischer-344 rats, is caused by differences in collateral blood flow rate through leptomeningeal anastomoses. In view of the retrograde direction of collateral blood flow into the middle cerebral artery territory, we developed parasagittal laser-Doppler flowmetry. Using this method two laser-Doppler probes are placed on the cerebral cortex: probe 1 is placed near the anastomoses between the middle- and anterior cerebral artery, probe 2 is placed 2 mm further away from these anastomoses than probe 1. We found in both rat strains a comparable relation between the areas under the curve of the signal measured by both laser-Doppler probes for 2 h after middle cerebral artery occlusion. This relation is considered to be a measurement of the collateral blood flow rate into the middle cerebral artery territory through leptomeningeal anastomoses after middle cerebral artery occlusion. We conclude that collateral blood flow for the two strains were essentially similar for the initial 2 h after MCA occlusion. Although these collateral blood flows could have been different at a later time, it is unlikely that the interstrain difference in cerebral infarction volume between Wistar and Fischer-344 rats after proximal middle cerebral artery occlusion is caused by an apparent interstrain difference in the magnitude of collateral blood flow rate through leptomeningeal anastomoses. The parasagittal laser-Doppler flowmetry technique we developed for these experiments is currently successfully used in our laboratory to evaluate the efficacy of hemodynamically active pharmacotherapeutical agents in raising the collateral blood flow rate into the middle cerebral artery territory after middle cerebral artery occlusion.

Distribution of Lys-gamma 2-melanocyte-stimulating hormone- (Lys-gamma 2-MSH)-like immunoreactivity in neuronal elements in the brain and peripheral tissues of the rat.

Using an antiserum raised against Lys- gamma 2-melanocyte-stimulating hormone (Lys- gamma 2-MSH), with a high specificity for this peptide and its des-Lys derivative, gamma 2-MSH, we found Lys- gamma 2-MSH-like immunoreactivity to have a widespread distribution in the rat brain. In colchicine-treated rats, groups of immunopositive cell bodies were found in the intermediate and anterior lobes of the pituitary gland, in the hypothalamic arcuate nucleus and in the commissural part of the nucleus of the solitary tract (NTS). Immunopositive fibers were found to originate from the latter two cell body regions. The distribution of these fibers was similar to that of the pro-opiomelanocortin containing cell bodies and projections as it has been described previously. Immunopositive terminals were found in brain region containing neurons which have been shown to express mRNA for melanocortin receptors, though the distribution of Lys-gamma 2-MSH-like immunoreactivity is considerably more widespread than that of mRNA for the 'gamma-MSH receptor' (the melanocortin MC3 receptor), which has been reported to be mainly expressed in the hypothalamus. In the periphery Lys-gamma 2-MSH immunoreactivity was localized in the adrenal medulla and in neuronal fibers and varicosities in the heart. The vascular system, the bronchi and kidney were immunonegative. The occurrence of Lys-gamma 2-MSH immunoreactivity in many of the brain regions which are involved in cardiovascular regulation offers leads for further studies on the putative role of gamma-MSHs in cardiovascular control. The occurrence in the rat heart of Lys-gamma 2-MSH-containing fibers suggests a role of the gamma-MSHs in cardiac function.

A comparative study on possible calcium antagonistic properties of indapamide and other drugs potentially interfering with calcium transport in isolated vascular smooth muscle.

In the present study the effects of indapamide (IN) were compared with those of chlorthalidone (C) and of drugs potentially interfering with calcium transport, e.g. verapamil (V), papaverine (P), phentolamine (PH) and diazoxide (D) using isolated rabbit aorta in order to detect calcium antagonistic properties. IN and C failed to show any relaxation effect towards either noradrenaline (NA)- or high K+-precontracted aorta strips, whereas V and P reduced the K+ contraction dose dependently and PH, V, P and D induced a significant relaxation of the NA contraction. Preincubation with IN or C did not affect the dose-response curve of NA-induced contractions. PH antagonized the NA contraction strongly whereas V and P shifted the dose-response curve to the right only at the highest concentration. In a Ca2+-depleted and high K+-depolarized aorta preincubation with V was able to antagonize Ca2+-induced contraction effectively in a dose-dependent fashion whereas P only showed partial inhibition at the highest concentration. The effects of the drugs on responses to NA in a Ca2+-free medium were also investigated. This type of contraction is likely to be due to mobilization of internally located Ca2+. The results demonstrate that PH was able to counteract this type of contraction effectively whereas only a high concentration of V, P and D was effective in reducing the Na contraction in Ca2+-free medium. Both IN and C failed to affect this type of contraction. In conclusion, the present results indicate that neither IN nor C possess calcium entry and or intracellular calcium antagonistic properties under different conditions as measured in a conduit vessel of rabbits. Moreover, the results point to an additional site of calcium antagonistic action for V, P and D, especially at higher concentrations.

The effects of gamma 2-melanocyte-stimulating hormone and nimodipine on cortical blood flow and infarction volume in two rat models of middle cerebral artery occlusion.

We observed that the pro-opiomelanocortin-derived neuropeptide, gamma 2-melanocyte-stimulating hormone (gamma 2-MSH), has various peripheral and central hemodynamic effects in the rat, including a marked enhancing effect on cerebral blood flow. This hemodynamic profile might be of interest in the pharmacotherapeutic approach to acute cerebral ischemia. Being an adrenocorticotropin (ACTH) analogue, gamma 2-MSH might also possess direct neuronal protective properties. Therefore, in two rat models of focal cerebral ischemia we studied the effects of gamma 2-MSH, with nimodipine, a Ca2+ channel antagonist, as a reference compound, on parasagittal laser-Doppler-assessed cortical blood flow and infarction volume. In isoflurane-anesthetized Wistar and F344 rats i.v. bolus infusions (four in total) of gamma 2-MSH or nimodipine or their vehicle controls were given 1 h before, 1 min after, and 1 h and 2 h after occlusion of the middle cerebral artery. We used both an intravasal and an extravasal middle cerebral artery occlusion technique because pilot experiments had shown differences in the severity of ischemia with the two techniques. gamma 2-MSH (100 nmol/kg in 1 min) increased cortical blood flow significantly but transiently, both pre- and post-ischemically, whereas nimodipine (20 micrograms/kg in 1 min) increased cortical blood flow only pre-ischemically in both models of middle cerebral artery occlusion. gamma 2-MSH had no effect on cortical and striatal infarction volume, while nimodipine caused a significant reduction of cortical infarction volume in the extravasal middle cerebral artery occlusion model. To conclude, despite its hemodynamic and possible neuroprotective properties, gamma 2-MSH did not prevent ischemic neuronal damage after middle cerebral artery occlusion in rats. This might be partly due to the short half-life of the peptide, leading to a transient increase in cortical blood flow and short neuronal exposure time, suggesting that prolonged infusion of the neuropeptide might be required. The results with nimodipine support the notion that it attenuates cortical ischemic damage, independently of effects on cerebral hemodynamics.

Disturbed adrenergic regulation of coronary flow in the guinea-pig heart after endotoxin.

We investigated the functional responsiveness of alpha- and beta-adrenoceptors in the isolated guinea-pig heart and coronary vascular system, 4 days after pretreatment with endotoxin. It was found that endotoxin reduced the flow-increasing effect of salbutamol and enhanced the flow-decreasing effect of B-HT 920. The positive chronotropic and inotropic effects of dobutamine and salbutamol and the flow-decreasing effect of phenylephrine were not influenced by endotoxin pretreatment. It is concluded that bacterial endotoxin induces alpha 2-adrenoceptor hyperreactivity and beta 2-adrenoceptor impairment in the coronary vascular system of the guinea-pig heart.

Further study of possible direct vascular actions of indapamide in the conduit and renal arteriolar vessels of spontaneously hypertensive rats.

The effect of indapamide on vascular reactivity and its properties as a calcium antagonist were studied in both isolated aorta and perfused renal vasculature of spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto (WKY) rats. Indapamide was given orally to SHR and WKY rats for 2 weeks at a dose of 5 mg/kg per day. During this period indapamide did not lower blood-pressure in SHR and WKY rats although there was an adequate concentration of indapamide in the blood. There were no differences observed in the vascular reactivity towards noradrenaline and high-K+ in both the above mentioned vessels in either indapamide- or vehicle-pretreated SHR and WKY rats. Verapamil (10(-9)-10(-5) M) caused a concentration-dependent relaxation of high-K+-depolarized aortas and a decrease in the renal-arteriolar perfusion pressure elevated by high-K+ in both strains of rat. However, indapamide (10(-7)-10(-4) M) did not affect the K+-induced effect on either vessel type. Preloading of the vessels in vivo with indapamide for 2 weeks did not influence the results. In conclusion, further evidence has been presented to show that indapamide does not have calcium-antagonist properties in conduit (aorta) or resistance (renal) vessels under hypertensive conditions. Preloading of the vessels with indapamide was not a prerequisite for the demonstration of a pharmacological action of indapamide.