Intrastriatal infusion of lisuride--a potential treatment for Parkinson's disease? Behavioral and autoradiographic studies in 6-OHDA lesioned rats

The present study examined the effect of chronic intrastriatal infusion of the dopamine receptor agonist lisuride on apomorphine-induced rotational behaviour and on D2-dopamine receptors in rats with unilateral 6-hydroxydopamine (6-OHDA) lesions of the nigrostriatal dopaminergic pathway. The completeness of the lesion of the right ascending nigrostriatal dopaminergic pathway was confirmed by apomorphine-induced rotation and [3H]-mazindol autoradiography. The intrastriatal infusion of lisuride (0.5 microgram/h) into the lesioned striatum for 2 weeks induced an immediate but temporary spontaneous contralateral rotation and a reduction of apomorphine-induced rotation of 47.2% relative to pre-lisuride infusion. The density of D2-receptors in the lisuride-infused striatum was significantly decreased by 40% relative to vehicle-infused 6-OHDA lesioned rats. The level of D2-dopamine receptors returned to normal levels 3 weeks after the termination of lisuride infusion. These results show that the intrastriatal infusion of lisuride reverses the behavioural and D2-dopamine receptor changes present in the 6-OHDA lesion rat model of Parkinson's disease.     

The role of nigrostriatal dopamine in metabotropic glutamate agonist-induced rotation

Metabotropic glutamate receptors are a major class of excitatory amino acid receptors. Eight metabotropic glutamate receptors subtypes have been cloned and have been classified into three groups based on their amino acid sequence homology, effector systems, and pharmacological profile. Previous results have shown that striatal group I metabotropic glutamate receptor stimulation produces vigorous contralateral rotation in intact rats, thought to be due to increased striatal dopamine release. Examination of FOS-like immunoreactivity and local cerebral glucose metabolism suggests that this occurs secondary to activation of the subthalamic nucleus. We sought to determine the contribution of dopamine by examining metabotropic glutamate receptor agonist-induced rotation in rats following acute dopamine depletion by reserpine/alpha-methyl-para-tyrosine treatment, or chronic dopamine depletion by 6-hydroxydopamine treatment. In unilateral 6-hydroxydopamine lesioned rats, the group I metabotropic glutamate receptor agonist (RS)-3,5-dihydroxyphenylglycine induced contralateral rotation with a coincident increase in striatal 3,4-dihydroxyphenylacetic acid. The rotation was attenuated by the group I antagonist 1-aminoindan-1,5-dicarboxylate. Examination of FOS-like immunoreactivity and [14C]2-deoxyglucose uptake in chronically dopamine depleted rats also revealed similar patterns to those seen previously in intact rats. However, acutely dopamine depleted rats do not exhibit metabotropic glutamate receptor agonist-induced rotation and show a different pattern of [14C]2-deoxyglucose uptake, with no increase in glucose utilization in the intermediate and deep layers of the superior colliculus. These results suggest that there are compensatory changes under conditions of chronic dopamine denervation which permit metabotropic glutamate receptor agonist-induced rotation to occur, which may include dopamine receptor supersensitivity, increased dopamine turnover, and/or changes in sensitivity of striatal group I metabotropic glutamate receptors. The group III metabotropic glutamate receptor agonist L-(+)-2-amino-4-phosphonobutyrate induced contralateral rotation in 6-hydroxydopamine lesioned rats, while it had no effect in intact rats. Additionally, examination of FOS-like immunoreactivity revealed a distinct pattern following L-(+)-2-amino-4-phosphonobutyrate administration in 6-hydroxydopamine lesioned versus intact rats. These results suggest that there is a change in the effect of striatal group III stimulation under conditions of dopamine depletion.     

Treadmill running induces striatal Fos expression via NMDA glutamate and dopamine receptors

Several non-physiological stimuli (i.e. pharmacological or electrical stimuli) have been shown to induce Fos expression in striatal neurons. In this work, striatal Fos (i.e. Fos-like) expression was studied after physiological stimulation, i.e. motor activity (treadmill running at 36 m/min for 20 min). In rats killed 2 h after the treadmill session, Fos expression was observed in the medial region of the rostral and central striatum, and in the dorsal region of the caudal striatum. Fos expression was prevented by pretreatment with the non-competitive N-methyl-D-aspartate (NMDA) glutamate receptor antagonist MK-801 (0.1 mg/kg) or the D1 dopamine receptor antagonist SCH-23390 (0.1 mg/kg), but not by pretreatment with the D2 receptor antagonist eticlopride (0.5 mg/kg). Thirty-six hours after 6-hydroxydopamine lesion, a considerable reduction in treadmill-induced Fos expression was observed in both sides; however, Fos expression in the lesioned striatum was higher than in the contralateral intact striatum. Several weeks after unilateral 6-hydroxydopamine lesion of the nigrostriatal system, treadmill-induced Fos expression was significantly, but not totally, reduced in the lesioned striatum. Corticostriatal deafferentation also led to considerable reduction in treadmill-induced Fos expression. The present results indicate that exercise induces striatal Fos expression and that, under physiological stimulation, concurrent activation of D1 and NMDA receptors is necessary for such expression to occur. Reduction of Fos expression is practically absolute after acute blockage of these receptors, but not after lesions, possibly due partially to compensatory changes.     

Binding of the nonpeptide antagonist, SB 209670, to endothelin receptors on cultured neurons

We studied the binding characteristics of a novel, nonpeptide endothelin antagonist, SB 209670, to two subtypes of endothelin (ET) receptor in cultured rat cerebellar granule cell neurons. Displacement binding studies of [125I]ET-1 performed in the presence of the ETB receptor-selective agonist, sarafotoxin 6c (S6c), allowed us to measure a Ki of 4.0 +/- 1.5 nM for (+/-)SB 209670 at the ETA receptor (n = 4). Similarly, binding studies in the presence of the ETA receptor-selective antagonist, BQ123, allowed us to measure a Ki of 46 +/- 14 nM for (+/-)SB 209670 at the ETB receptor (n = 4). These studies indicate that the novel endothelin antagonist, SB 209670, has high affinity for both types of neuronal endothelin receptor.     

Margatoxin increases dopamine release in rat striatum via voltage-gated K+ channels

The distribution of iodinated margatoxin ([125I]margatoxin) binding sites in rat was investigated by autoradiography. Rat striatum expresses a high density of margatoxin binding sites and, therefore, the effects of margatoxin, charybdotoxin and iberiotoxin have been studied on [3H]dopamine release from rat striatal slices in vitro. Margatoxin (0.1-100 nM) and charybdotoxin (10-1000 nM), but not iberiotoxin increased the spontaneous and the electrically evoked [3H]dopamine release. [3H]dopamine release by margatoxin was inhibited by tetrodotoxin and omega-conotoxin GVIA, but not by atropine, naloxone, N(omega)-nitro-L-arginine and neurokinin or neurotensin receptor antagonists. In the buffer solution used for release experiments, [125I]margatoxin labels a maximum of 0.12 pmol of sites/mg protein in rat striatal membranes with a Kd of 5 pM. [125I]margatoxin binding was inhibited by margatoxin (Ki of 4 pM), charybdotoxin (Ki of 162 pM) but not by iberiotoxin. We conclude that inhibition of margatoxin-sensitive voltage-gated K+ channels increases [3H]dopamine release demonstrating their role in repolarization of nigrostriatal projections. In contrast, iberiotoxin-sensitive, high-conductance Ca2+-activated K+ channels are not involved in release of [3H]dopamine.     

Nonpeptide endothelin receptor antagonists. XI. Pharmacological characterization of SB 234551, a high-affinity and selective nonpeptide ETA receptor antagonist

The present study describes the pharmacological profile of ((E)-alpha-[[1-butyl-5-[2-[(2-carboxyphenyl)methoxy]-4-methoxy-phenyl ]-1H-pyrazol-4-yl]methlene]-6-methoxy-1,3-benzodioxole-5-propanoic acid) (SB 234551), a high-affinity, nonpeptide endothelin type A (ETA)-selective receptor antagonist. In human cloned ETA and endothelin type B (ETB) receptors, SB 234551 produced a concentration-dependent displacement of [125I]-endothelin-1 with Ki values of 0.13 and 500 nM, respectively. SB 234551 elicited concentration-dependent, rightward competitive shifts in the endothelin-1 concentration-response curves in isolated rat aorta and isolated human pulmonary artery (ETA receptor-mediated vascular contraction) with Kb values of 1.9 and 1.0 nM, respectively. SB 234551 antagonized ETB receptor-mediated vasoconstriction in the isolated rabbit pulmonary artery, as demonstrated by concentration-dependent, rightward shifts in the sarafotoxin S6c concentration-response curves (Kb = 555 nM). SB 234551 produced weak functional inhibition of sarafotoxin S6c-mediated endothelium-dependent relaxation (IC50 = 7 microM). SB 234551 (10 microM) had no significant effect against contraction produced by several other vasoactive agents and did not significantly influence radioligand binding to a number of diverse receptors. SB 234551 (0. 1-1.0 mg/kg i.v.) dose-dependently inhibited the pressor response to exogenous endothelin-1 in conscious rats. In vivo pharmacokinetic analysis in the rat demonstrated that SB 234551 was rapidly absorbed from the GI tract with a bioavailability of 30%. SB 234551 had a plasma half-life of 125 min and a systemic clearance of 25.0 ml/min/kg. The present study demonstrates that SB 234551 is an antagonist with high affinity for the ETA receptor, while sparing the ETB receptor. SB 234551 is a new pharmacological tool that should assist in the elucidation of the role of endothelin in pathophysiology.     

Atypical receptors mediate the response to endothelin-1 and sarafotoxin S6b in the human umbilical artery

The receptors mediating smooth muscle response to endothelin-1 and sarafotoxin S6b in the human umbilical artery were investigated in vitro. Both agonists induced contractions that were unaffected by the endothelin ET(B) receptor antagonist BQ 788 (10(-9), 10(-8), 10(-7) M). The non-selective endothelin ET(A/B) receptor antagonist PD 142893 (10(-7) M) decreased the contraction induced by endothelin-1. PD 142893 (10(-9) M) enhanced the contraction induced by sarafotoxin S6b whereas higher concentrations had no effect. Removing the endothelium did not affect the antagonising action of PD 142893 on endothelin-1-induced contractions while the enhancement of the sarafotoxin S6b-induced contraction was abolished. Sarafotoxin S6b induced relaxation in segments precontracted by 5-hydroxytryptamine and exposed to the endothelin ET(A) receptor antagonist BQ 123 (10(-7) M) and PD 142893 (10(-9) M) abolished this relaxation. These endothelial receptors seem neither to be classical endothelin ET(A) nor endothelin ET(B) receptors and they are not activated by endothelin-1.     

The mechanism of myometrial contractions induced by endothelin-1 in rat

Experiments were performed to characterize endothelin-1-induced contractions and the role of endothelin (ET) receptor subtypes in rat myometrium. The binding sites of [(125)I]-ET-1 were saturable with high affinity. Scatchard plot analysis revealed that ET-1 binding sites in the myometrium constituted a single population. The dissociation equilibrium constant (Kd) and the maximum binding sites (Bmax) were determined to be 48.9+/-3.0 pM and 1364.0+/-210.3 fmol/mg protein respectively. Specific [(125)I]-ET-1 binding was inhibited completely by unlabelled ET-1 and Ro 46-2005 (mixed-type ET receptor antagonist), but not fully (90.7+/-1.4%) by BQ 123 (a selective ETA receptor antagonist), and not at all by RES 701-1 (a selective ETB receptor antagonist). ET-1 induced myometrial contractions were composed of two types, an increase in resting tone and rhythmic contractions. These contractions were inhibited by BQ 123 and Ro 46-2005, but not by RES 701-1. ET-1-induced contractions were greatly reduced in Ca2+-free Krebs' solution. Nifedipine abolished the rhythmic contractions without affecting the increase in resting tone. These results suggest that ETA receptors are predominantly localized in rat myometrium and that excitation of ETA receptors evokes two types of contractions by increasing the cytoplasmic Ca2+ concentration.     

Endothelin ET(B) receptor-mediated constriction in the rabbit basilar artery

The present study tests whether endothelin ET(B) receptor activation can mediate endothelin-1 constriction in the rabbit basilar artery in situ. Endothelin-1 (30 nM) induced 27% constriction of vessels pretreated with 1 microM BQ610 (homopiperidenyl-CO-Leu-DTrp (CHO)-D-Trp-OH), an endothelin ET(A) receptor antagonist, and the resulting constriction was completely relaxed by BQ788 (N-cis-2,6-dimethylpiperidinocarbonyl L-gamma-MeLeu-D-Trp (COOCH3)-Nle), an endothelin ET(B) receptor antagonist. Similarly, 30 nM endothelin-1 induced 30% constriction of vessels pretreated with 1 microM BQ788, and the resulting constriction was completely relaxed by BQ610. In contrast, sarafotoxin S6c, an endothelin ET(B) receptor agonist, did not induce constriction. This study suggests that in the basilar artery (1) endothelin ET(B) receptor activation can result in constriction and (2) the ability to elicit constriction is in some way dependent upon the agonist that activates the endothelin ET(B) receptor.     

Motor stimulant effects of caffeine in 6-hydroxydopamine-lesioned rats are dependent on previous stimulation of dopamine receptors: a different role of D1 and D2 receptors

Caffeine has been reported to induce contralateral rotational behaviour in rats bearing a unilateral 6-hydroxydopamine lesion of the dopaminergic nigrostriatal pathway. In order to define the role of dopamine receptors in the mediation of this behaviour, we have evaluated the influence of previous exposure to a dopamine receptor agonist and the importance of the time elapsed from the 6-hydroxydopamine lesion on the rotational behaviour induced by caffeine. Separate groups of rats lesioned with 6-hydroxydopamine 2 weeks previously were exposed to four administrations of the D1/D2 receptor agonist apomorphine (0.3 mg/kg s.c.) (primed) or vehicle (drug-naive). Three days later, all rats received caffeine (30 mg/kg s.c.). Drug-naive 6-hydroxydopamine-lesioned rats did not rotate in response to caffeine, while rats primed with apomorphine rotate contralaterally in response to caffeine. When apomorphine priming was paired to the same environment (hemispherical bowls) where rats received caffeine, rotational behaviour was significantly higher than that obtained in rats primed in an unpaired environment (cylinders). Repeated priming with the D2/D3 receptor agonist quinpirole (0.2 mg/kg s.c.) induced a totally context-dependent contralateral rotation in response to caffeine, while caffeine contralateral rotation was not dependent from the context after repeated priming with the D1 agonist SKF 38393 [1-phenyl-2,3,4,5-tetrahydro-(1 H)-3-benzazepine-7,8-diol hydrochloride, 3 mg/kg s.c.]. Caffeine-mediated contralateral rotation was also evaluated in rats lesioned with 6-hydroxydopamine 12 weeks previously and exposed to four administrations of apomorphine or vehicle. As for rats repeatedly exposed to vehicle or apomorphine 2 weeks after 6-hydroxydopamine lesioning, caffeine failed to induce contralateral rotation in drug-naive rats, while it did induce a partially context-dependent contralateral rotation in apomorphine-primed rats. Different from rats receiving apomorphine priming 2 weeks after 6-hydroxydopamine lesioning, in 12 week-lesioned rats, caffeine also induced contralateral rotation after one priming with apomorphine (0.3 mg/kg s.c.), a condition which fails to induce context-dependent rotation. Administration of selective antagonists of A1 (8-cyclopentyl-1,3-dipropylxanthine), (DPCPX) or A2A (5-amino-2-(2-furyl)-7-(3-phenylpropyl)-pyrazolo[4,3-e]-1 ,2,4-triazolo[5c]pirimidine), (SCH 58261) adenosine receptors failed to induce contralateral rotation either alone or in combination in 12 week-6-hydroxydopamine-lesioned rats repeatedly primed with apomorphine. All together, the results indicate that: (i) caffeine does not induce any contralateral rotation in drug-naive 6-hydroxydopamine-lesioned rats; (ii) priming with a dopamine agonist enables caffeine to induce contralateral rotation, this rotation is, however, context independent only after priming with a selective D1 agonist; (iii) contralateral rotation in response to caffeine is dependent on the time from the 6-hydroxydopamine lesion; (iv) blockade of A1 and A2A adenosine receptors with selective antagonists does not induce contralateral rotational behaviour in 6-hydroxydopamine-lesioned rats.     

Endothelin-1-(1-31), a novel vasoactive peptide, increases [Ca2+]i in human coronary artery smooth muscle cells

We have previously found that human chymase cleaves big endothelins at the Tyr31-Gly32 bond and produces 31-amino acid long endothelins-(1-31), without any further degradation products. In this study, we investigated the effect of synthetic endothelin-1-(1-31) on the intracellular free Ca2+ concentration ([Ca2+]i) in cultured human coronary artery smooth muscle cells. Endothelin-1-(1-31) increased [Ca2+]i in a concentration-dependent manner (10(-14) to 10(-10) M). This endothelin-1-(1-31)-induced [Ca2+]i increase was not affected by phosphoramidon (N-(alpha-Rhamnopyranosyloxyhydroxyphosphinyl)-L-Leucyl-L-Tryptoph an), an inhibitor of endothelin-converting enzyme. It was, however, inhibited by 10(-10) M BQ123 (Cyclo-(-D-Trp-D-Asp(ONa)-Pro-D-Val-Leu-)), an endothelin ET(A) receptor antagonist, but not by 10(-10) M BQ788 (N-cis-2,6-dimethylpiperidinocarbonyl-L-yMeLeu-D-Trp(COOM e)-D-Nle-ONa), an endothelin ET(B) receptor antagonist. These results suggest that endothelin-1-(1-31) by itself exhibits vasoactive properties probably through endothelin ET(A) receptors. Since human chymase has been reported to play a role in atherosclerosis, endothelin-1-(1-31) may be one of the candidate substances for its cause.     

Decline in striatal dopamine D1 and D2 receptor activation in aged F344 rats

Aging differentially affects receptor function. In the present electrophysiological study we compared neuronal responsiveness to locally applied dopamine D1 and D2 receptor agonist in the striatum of female Fischer 344 rats aged 3 and 26-27 months. In a subgroup of the old rats, the nigrostriatal dopamine bundle was destroyed unilaterally with 6-hydroxydopamine (6-OHDA) to assess receptor plasticity in response to denervation. Spontaneous firing rate of striatal neurons was higher in aged compared to young rats. Higher doses of the D1 agonist SKF 38393 or the D2 agonist quinpirole were required to elicit a 50% change in firing rate in aged compared to young rats. No difference with SKF 38393 or quinpirole was detected between 6-OHDA denervated and control (nonlesioned) striatum in aged rats. Supersensitivity to D2 agonists has been reported following 6-OHDA lesions in young rats. These observations suggest that D2 receptors in aged rat striatum might not be as plastic as in younger rats.     

Mechanisms of endothelin-induced macromolecular leakage in microvascular beds of rat mesentery

Microvascular responses to endothelin-3 were investigated in the rat mesentery under fluorescence microscopy. Endothelin-3 in a range of 0.1-100 pM induced arteriolar constriction in a dose-dependent manner, and stimulated Ca2+ mobilization, demonstrated by fura-2-associated fluorography, in both arterioles and venules. Cyclo(-D-Trp-D-Asp-Pro-D-Val-Leu-) (BQ123), and endothelin ETA receptor antagonist, at a concentration of 10 microM inhibited the endothelin-3 (100 pM)-induced arteriolar constriction and Ca2+ mobilization in arterioles but not in venules. In venules, an early onset leakage of FITC (fluorescein isothiocyanate)-labeled albumin and subsequent reduction of red blood cell velocity without arteriolar constriction were observed after the superfusion of endothelin-3 with BQ123, suggesting that a non-endothelin ETA receptor mediates macromolecular leakage followed by a decrease in blood flow. Endothelin-3 with BQ123 neither stimulated leukocyte adhesion nor activated luminol-dependent chemiluminescence in venules, showing that endothelin-3-increased permeability may be induced by leukocyte-independent and oxyradical-independent mechanisms. These microvascular alterations of permeability and red blood cell velocity were significantly attenuated by the addition of phalloidin, an F-actin stabilizer, suggesting the involvement of endothelial cell contraction. Nicardipine (1,4-dihydro-2,6-dimethyl-4-[3-nitrophenyl]methyl-2- [methyl(phenylmethyl)amino]-3,5-pyridinedicarboxylic acid ethyl ester), a dihydropyridine-type Ca2+ channel antagonist, eliminated endothelin-3-induced arteriolar constriction; however, it did not affect albumin leakage promoted by endothelin-3 with BQ123, suggesting that a non-voltage-dependent Ca2+ channel(s) is involved in non-endothelin ETA receptor-mediated Ca2+ mobilization and contraction of venular endothelial cells. Overall, it is conceivable that endothelin ETA receptor and voltage-dependent Ca2+ channel are involved in endothelin-3-induced arteriolar constriction. In addition, the present results suggest that Ca2+ mobilization in venular endothelium, which is mediated by a non-endothelin ETA receptor, possibly endothelin ETB receptor and regulated by non-voltage-dependent Ca2+ channel(s), may cause endothelial cell contraction and subsequently increase macromolecular permeability in microvascular beds treated with endothelin-3.     

Effects of Ro 47-0203 and PD155080 on the plasma kinetics, receptor binding and vascular effects of endothelin in the pig

The effects of the mixed endothelin ET(A)/endothelin ET(B) receptor antagonist Ro 47-0203 (bosentan, 4-tert-butyl-N-[6-(2-hydroxy-ethoxy)-5-( 2-methoxy -phenoxy)-2,2'-bipyrimidin-4-yl] -benzenesulfonamide) and the selective endothelin ET(A) receptor antagonist PD155080 (sodium 2-benzo[1,3]dioxol-5-yl-3-benzyl-4-(4-me thoxy-phenyl)-4-oxobut+ ++-2-enoate) on plasma half-life and regional extraction of exogenous endothelin-1 as well as on the regional vascular effects of endothelin-1 were investigated in the pig in vivo. Bosentan but not PD155080 (5 mg/kg, i.v. bolus, both drugs) increased the arterial plasma levels of endothelin-1-like immunoreactivity. Neither of the drugs affected the plasma half-life of infused endothelin-1. In the spleen, both the extraction and vascular effects of exogenous endothelin-1 were attenuated by both bosentan and PD155080 whereas renal extraction and vascular effects in the kidney were unaffected by both drugs. In the lung, only bosentan decreased pulmonary extraction of endothelin-1. In conclusion, the bosentan-induced increase of circulating endothelin-1 seems to be related to blockade of endothelin-1 binding to endothelin ET(B) receptors. Blockade of these receptors does not influence the overall elimination of endothelin-1, however.     

Ethanol inhibits mitogen activated protein kinase activity and growth of vascular smooth muscle cells in vitro

The intrathecal (i.t.) injection of endothelins to conscious rats was found to cause respiratory arrest. To gain some insights into this central phenomenon, peripheral vascular permeability and lung oedema were measured after i.t. and i.v. injections of these peptides. When injected at T-8 spinal cord level, endothelin-1 (65 and 650 pmol) and endothelin-3 (650 pmol) enhanced vascular permeability in the lungs by 22-fold and 7-fold, respectively, and caused sudden death at the highest dose. Less prominent increases (between 1.4- and 2.2-fold) of vascular permeability were observed in other tissues (trachea, kidney, ears, skin of hind paws and back skin) with endothelin-1. Endothelin-1 (650 pmol) caused a similar increase (27-fold) in lung vascular permeability when injected at T-2, although the response was significantly less (P < 0.05) if injected at the L-4 (15-fold) spinal cord level. Only endothelin-1 produced lung oedema when injected at the T-2 or T-8 level. In contrast, intravenous injection of endothelins-1 and -3 (650 pmol) did not produce lung oedema and the lung vascular permeability was increased by only 1.4-1.6-fold and all rats survived. The prior i.t. injection of 6.5 nmol BQ-123 (cyclo[D-Trp, D-Asp, L-Pro, D-Val, L-Leu]), a selective endothelin ET(A) receptor antagonist, prevented the increases of lung vascular permeability and oedema and the mortality induced by i.t. endothelin-1 (650 pmol). Whereas i.v. treatment with phentolamine (2 mg/kg) or pentolinium (25 mg/kg + 50 mg/kg per h x 15 min) abolished the lung vascular permeability changes evoked by endothelin-1 (650) pmol), atropine (1 mg/kg), NG-nitro-L-arginine (50 mg/kg) or indomethacin (5 mg/kg) had no effect. Moreover, the effects of endothelin-1 were attenuated in capsaicin pretreated rats (125 mg/kg, 10 days earlier) and almost abolished in rats subjected to sympathectomy with 6-hydroxydopamine (100 mg/kg, 24-48 h earlier). All these treatments except atropine and NG-nitro-L-arginine prevented the endothelin-1-induced lung oedema and reduced the lethality by around 50%. These results suggest that the increases of pulmonary vascular permeability and oedema induced by i.t. endothelin-1 are due to an intense pulmonary vasoconstriction mediated by alpha-adrenoceptors following the release of catecholamines in response to the activation of endothelin ET(A) receptor in the spinal cord. This central phenomenon seems to be reflexogenic, including the involvement of primary afferent C-fibers and spinal cord ascending fibers to the brain. Thus, endothelin-1 could play a role in neurogenic pulmonary oedema through a central mechanism.     

Localization of serotonin-4 receptors in the striatonigral pathway in rat brain

Rats were injected unilaterally with 6-hydroxydopamine either in the medial forebrain bundle or in the dorsolateral substantia nigra. Another group was injected unilaterally with kainate in the striatum. The loss of neurons was assessed by a reduction in tyrosine hydroxylase-like immunoreactivity for dopaminergic neurons, and choline acetyltransferase-like and glutamate decarboxylase-like immunoreactivities for cholinergic and GABAergic neurons, respectively. Brain sections also were analysed by autoradiography on 20 micron sections with the radio-iodinated serotonin-4 receptor antagonist [125I]SB 207710 [Brown A. M. et al. (1993) Br. J. Pharmac. 110, 10P]. Kainate injections in the striatum resulted in loss of choline acetyltransferase- and glutamate decarboxylase-like immunoreactive cell bodies in this area. There was also a decrease in glutamate decarboxylase-like immunoreactivity on the ipsilateral side in the substantia nigra and entopeduncular nucleus. These changes were accompanied by substantial (> 50%) decreases in [125I]SB 207710 binding in both the ipsilateral striatum (confined to the lesioned area) and substantia nigra, with no change in either the nucleus accumbens or the globus pallidus. There was also significant loss of [125I]SB 207710 binding in the ipsilateral entopeduncular nucleus. 6-Hydroxydopamine lesions placed either in the medial forebrain bundle or in the substantia nigra failed to decrease [125I]SB 207710 binding in any of these areas, although there was total loss of tyrosine hydroxylase-like immunoreactive terminals in the striatum and cell bodies in the nigra. We conclude that serotonin-4 receptors are present on projection neurons, both on their perikarya in the striatum and terminals in the nigra and entopeduncular nucleus. It is likely that these receptors are located on the GABAergic projection neurons and possibly on cholinergic and GABAergic interneurons. However, serotonin-4 receptors are not located on dopaminergic neurons, either on their cell bodies in the substantia nigra or terminals in the striatum.     

Endothelin-1 induces vasoconstriction and nitric oxide release via endothelin ET(B) receptors in isolated perfused rat liver

Endothelin-1 (0.1, 1 and 10 nM) induced a significant increase in portal pressure and nitric oxide (NO) release in the isolated rat liver. The endothelin ET(B) receptor agonist, IRL 1620 (Suc-[Glu9,Ala(11,15)]endothelin-1-(8-21)) (0.1, 1 and 10 nM) also elicited a marked increase in portal pressure and NO release. The potency of endothelin-1 was higher than that of IRL 1620. The endothelin ET(A) receptor antagonist, BQ-123 (cyclo(-D-Trp-D-Asp-Pro-D-Val-Leu)) (1 and 10 microM), had no effect on the endothelin-1-induced change in portal pressure and NO current. In contrast, the endothelin ET(B) receptor antagonist, BQ-788 (N-cis-2,6-dimethylpiperidinocarbonyl-L-gamma-methyl-leucyl-D-1-++ +methoxycarbonyltryptophanyl-D-norleucine) (1 and 10 nM), attenuated the endothelin-1-induced change in portal pressure and NO current. Administration of N(G)-monomethyl-L-arginine (L-NMMA), a NO synthase inhibitor, completely abolished the endothelin-1- or IRL 1620-induced NO release. L-NMMA enhanced the increase in portal pressure and decrease in O2 consumption caused by endothelin-1. These results indicated that endothelin ET(B) receptors mediate both vasoconstriction and NO release and that NO plays a significant role in stabilizing microcirculation in isolated perfused rat liver.     

Systemic morphine-induced Fos protein in the rat striatum and nucleus accumbens is regulated by mu opioid receptors in the substantia nigra and ventral tegmental area

To characterize how systemic morphine induces Fos protein in dorsomedial striatum and nucleus accumbens (NAc), we examined the role of receptors in striatum, substantia nigra (SN), and ventral tegmental area (VTA). Morphine injected into medial SN or into VTA of awake rats induced Fos in neurons in ipsilateral dorsomedial striatum and NAc. Morphine injected into lateral SN induced Fos in dorsolateral striatum and globus pallidus. The morphine infusions produced contralateral turning that was most prominent after lateral SN injections. Intranigral injections of [D-Ala2, N-Me-Phe4, Gly-ol5]-enkephalin (DAMGO), a mu opioid receptor agonist, and of bicuculline, a GABAA receptor antagonist, induced Fos in ipsilateral striatum. Fos induction in dorsomedial striatum produced by systemic administration of morphine was blocked by (1) SN and VTA injections of the mu1 opioid antagonist naloxonazine and (2) striatal injections of either MK 801, an NMDA glutamate receptor antagonist, or SCH 23390, a D1 dopamine receptor antagonist. Fos induction in dorsomedial striatum and NAc after systemic administration of morphine seems to be mediated by dopamine neurons in medial SN and VTA that project to medial striatum and NAc, respectively. Systemic morphine is proposed to act on mu opioid receptors located on GABAergic interneurons in medial SN and VTA. Inhibition of these GABA interneurons disinhibits medial SN and VTA dopamine neurons, producing dopamine release in medial striatum and NAc. This activates D1 dopamine receptors and coupled with the coactivation of NMDA receptors possibly from cortical glutamate input induces Fos in striatal and NAc neurons. The modulation of target gene expression by Fos could influence addictive behavioral responses to opiates.     

Dopamine receptors status after unilateral nigral 6-OHDA lesion. Autoradiographic and in situ hybridization study in the rat brain

The physiological effects of dopamine (DA) are mediated by several distinct receptor subtypes. The effects of unilateral nigral 6-hydroxydopamine (6-OHDA) lesions on DA receptors were investigated by receptor autoradiography using the D1 selective ligand [3H]SCH 23390 as well as the D2 ligand [3H]spiroperidol. mRNA distribution was studied by in situ hybridization. Lesioned rats were sacrificed at different time intervals. Receptor binding studies were performed on tissue sections using selective ligands. [35S]UTP labeled RNA probes were prepared from the different cDNA (D1, D2, D3) and used for in situ hybridization. A specific loss of receptor binding sites and mRNA hybridization was found in the lesioned substantia nigra pars compacta (SNc) at all times examined. Receptor binding studies revealed a different time-dependent increase in both D1 and D2 receptors. In situ hybridization showed that only D2 receptor mRNA increased in the caudate-putamen (CPu) of the lesioned side 15 d after 6-OHDA. No changes were observed in D1 and D3 receptor mRNA during the entire time-course.