The effect of cyclo (Leu-Gly) on chemical denervation supersensitivity of dopamine receptors induced by intracerebroventricular injection of 6-hydroxydopamine in mice

Chemical denervation supersensitivity of postsynaptic dopamine receptors was induced in mice by intracerebroventricular injection of 6-hydroxydopamine. Fourteen days after the 6-hydroxydopamine injection, mice exhibited greater spontaneous locomotor activity and hypothermic response when challenged intraperitoneally with apomorphine. Whole brain levels of dopamine were reduced by 80%. Daily subcutaneous administration of cyclo (Leu-Gly) (50 μg/mouse/day) for 14 days inhibited the development of dopamine receptor supersensitivity induced by 6-hydroxydopamine as evidenced by the blockade of an apomorphine induced locomotor and hypothermic effect. Cyclo (Leu-Gly) did not alter the depletion of brain dopamine induced by 6-hydroxydopamine. These data suggest that cyclo (Leu-Gly) can block the development of dopamine receptor supersensitivity induced by 6-hydroxydopamine without protecting the neurons from dopamine depletion.     

Effects of cyclo (Leu-Gly) on neurochemical indices of striatal dopaminergic supersensitivity induced by prolonged haloperidol treatment

The effects of a prolonged treatment with cyclo (Leu-Gly) and/or haloperidol on biochemical parameters indicative of striatal dopamine target cell supersensitivity have been investigated in the rat. When given acutely, cyclo (Leu-Gly) (2 mg/kg sc) did not affect striatal homovanillic acid, dihydroxyphenylacetic acid and acetylcholine levels both under basal conditions or after acute haloperidol (1 mg/kg ip) treatment. When given concomitantly with haloperidol (infused by means of osmotic minipumps at a rate of 2.5 μg/h sc) for 14 days, cyclo (Leu-Gly) (2 mg/kg sc once daily) failed to prevent the fall of striatal dopamine metabolites observed 2 days following withdrawal and the tolerance to the elevation of dopamine metabolites which occurs in response to challenge with the neuroleptic during withdrawal. Prolonged treatment with cyclo (Leu-Gly) also failed to affect the tolerance to the decrease of striatal acetylcholine levels which occurs under chronic haloperidol treatment. These data suggest that the mechanism whereby cyclo (Leu-Gly) inhibits the development of neuroleptic-induced dopaminergic supersensitivity does not involve an action of the peptide on nigro-striatal dopaminergic and striatal cholinergic neurons and is probably exerted distally to both dopaminergic and cholinergic synapses.     

Dihydroergotoxine decreases blood pressure in spontaneously hypertensive rats by interacting with peripheral dopamine receptors

Dihydroergotoxine (10 micrograms/kg s.c.) decreased mean carotid blood pressure in urethane-anaesthetized spontaneously hypertensive rats but failed to modify the same parameter in normotensive rats. The effect was statistically significant 20 min after the injection and relatively long lasting (up to 90 min). Pharmacological characterization of the phenomenon indicated that it is mediated by stimulation of dopamine receptors, since pretreatment with haloperidol, cis-flupentixol but not with trans-flupentixol, completely prevent the reduction in blood pressure induced by dihydroergotoxine. Moreover, a challenge dose of dihydroergotoxine did not reduce mean blood pressure values in spontaneously hypertensive rats pretreated with domperidone or (-)sulpiride, but not with (+)sulpiride. These results suggest that the ergot derivative modifies the cardiovascular system by interaction with peripheral dopamine receptors of the DA2 type.     

Antagonism of kappa opioid mediated effects in the rat by cyclo(Leu-Gly)

The effect of cyclo(Leu-Gly) on U-50,488H- induced pharmacological actions was determined in male Sprague-Dawley rats. Intraperitoneal (i.p.) administration of U-50,488H to rats produced analgesia (tail-flick) and increased urinary output. Cyclo (Leu-Gly) (1-4 mg/kg, s.c.) antagonized the analgesic response to U-50,488H (25 mg/kg; i.p.). A dose of 10 mg/kg (i.p.) of U-50,488H increased the spontaneous urinary output which was antagonized by cyclo (Leu-Gly) (1-4 mg/kg; s.c.). To determine whether cyclo (Leu-Gly) was acting as a kappa-opioid receptor antagonist, the effect of cyclo (Leu-Gly) on the binding of [3H]ethylketocyclazocine (EKC) to membranes of rat cerebral cortex and spinal cord was determined. The IC50 values of cyclo(Leu-Gly) in displacing [3H]EKC from its binding sites in cortex and spinal cord were 1.44 and 0.40 mM, respectively. Chronic administration of U-50,488H (25 mg/kg; i.p., b.i.d.) for 4 days induced tolerance to its analgesic effect. The latter was not affected by cyclo(Leu-Gly) (2 to 8 mg/kg; s.c.) given once a day for 4 days. It is concluded that cyclo(Leu-Gly) antagonizes acute actions of U-50,488H and that such effects of cyclo(Leu-Gly) are not mediated via a direct action on kappa-opioid receptors.     

Effects of Pro-Leu-Gly-NH2 and cyclo (Leu-Gly) on the binding of 3H-quinuclidinyl benzilate to striatal cholinergic muscarinic receptors

The effects of Pro-Leu-Gly-NH2 (melanotropin release inhibiting factor, MIF) and its analog, cyclo (Leu-Gly) on the mouse and rat striatal cholinergic muscarinic receptors labeled with 3H-quinuclidinyl benzilate (QNB) were investigated. 3H-QNB bound to the rat striatal muscarinic receptors at a single high affinity site with receptor density (Bmax value) of 1200 fmol per mg protein and an apparent dissociation constant (Kd value) of 53.5 pM. At 140 pM concentration of 3H-QNB, the specific binding to the receptors was 724 fmol per mg protein. MIF in a concentration range of 10(-9) to 10(-4) M did not alter the binding of 3H-QNB but at 10(-3) M decreased the binding by 25%. Cyclo (Leu-Gly), on the other hand, in the concentration range of 10(-9) to 10(-3) M had no effect on the binding of 3H-QNB. A single injection of MIF (3 or 10 mg/kg IP) to rats did not alter the Bmax or the Kd value of 3H-QNB to bind to the striatal membranes. 3H-QNB bound to the mouse striatal muscarinic receptors at a single high affinity site with a Bmax value of 991 fmol/per mg protein and a Kd value of 21 pM. Neither acute administration of MIF (3 or 10 mg/kg IP) nor chronic treatment of the peptide (2, 8 or 32 mg/kg IP, daily for 5 days) to mice could influence the binding of 3H-QNB to the striatal muscarinic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cyclo (Leu-Gly) + haloperidol: effects on dopamine receptors and conditioned avoidance responding

Behavioral effects of cyclo (Leu-Gly) (cLG), administered either acutely or chronically, were assessed in combination with haloperidol in the rat. cLG administered chronically, produced a significant reduction in the increase in apomorphine-induced stereotypy produced by chronic haloperidol infusion. On the other hand, the same dose of cLG which reduced this induction of dopamine receptor supersensitivity due to chronic haloperidol treatment, failed to produce a change in the potency of haloperidol in blocking conditioned avoidance responding in the rat. Furthermore, degeneration-induced supersensitivity of dopamine neurons, produced by unilateral destruction of the nigrostriatal pathway, was not reduced by acute or chronic treatment with cLG as measured by apomorphine-induced rotation. These data suggest that cLG may decrease motor system side effects thought to be caused by chronic antipsychotic administration without affecting the therapeutic efficacy of the antipsychotic agent.     

Cyclo(Leu-Gly) has opposite effects on D-2 dopamine receptors in different brain areas

Cyclo(Leu-Gly) (cLG), a diketopiperazine analog of Pro-Leu-Gly-NH2 (MIF), affects a number of physiological and behavioral responses to the endogenous neurotransmitter, dopamine (DA). In the present series of experiments, the effect of in vivo administration of cLG (8 mg/kg) was investigated five days following subcutaneous administration. It was found that cLG administration of cLG (8 mg/kg) was investigated five days following subcutaneous administration. It was found that cLG administration caused a supersensitive behavioral response, measured by increased stereotypic sniffing, to the DA agonist, apomorphine (APO). At the same time, an increase was found in the affinity for dopamine (DA), as measured by dopamine inhibition of 3H-spiroperidol binding to D-2 DA receptors in striatum (nigro-striatal DA tract). In contrast, the same peptide treatment caused a subsensitive physiological response to APO-induced hypothermia, concomitant with a decrease in affinity for dopamine, as measured by DA inhibition of 3H-spiroperidol binding to D-2 DA receptors in hypothalamus (incerto-hypothalamic DA tract). These results suggest that a single neuromodulatory agent, the peptide cLG, can elicit diametrically opposite effects on D-2 DA receptors and on the corresponding physiological endpoints in two different brain areas.     

D(1) dopamine receptor regulation of NHE3 during development in spontaneously hypertensive rats

To determine if the defective interactions among D(1)-like receptors, G proteins, and Na(+)/H(+) exchanger 3 (NHE3) are consequences of hypertension, we studied these interactions in rats, before (2--3 wk) and after (12 wk) the establishment of hypertension. To eliminate the confounding influence of second messenger action on D(1) receptor-NHE3 interaction, studies were performed in renal brush-border membranes (BBM) devoid of cytoplasmic second messengers. NHE3 activity increased with age in Wistar-Kyoto (WKY) rats (3 wk = 1.48 +/- 0.39, n = 13; 12 wk = 2.83 +/- 0.15, n = 16, P < 0.05) but not in spontaneously hypertensive rats (SHRs; 3 wk = 2.52 +/- 0.37, n = 11; 12 wk = 2.81 +/- 0.20, n = 16). D(1) receptor protein tended to decrease, whereas NHE3 protein tended to increase with age in both WKY and SHRs. However, the inhibitory effect of a D(1)-like agonist, SKF-81297, on NHE3 activity increased with age in WKY rats (3 wk = -40.7 +/- 5.3%, n = 10, 12 wk = -58.7 +/- 4.6%, n = 12, P < 0.05) but not in SHRs (3 wk = -27.6 +/- 5.9%, n = 11, 12 wk = -25.1 +/- 3.2%, n = 11). The decreased inhibitory effect of another D(1)-like agonist, fenoldopam, on NHE3 activity in SHRs was not caused by increased activity and binding of G beta gamma to NHE3 as has been reported in young WKY rats. G(s)alpha mediates, in part, the inhibitory effect of D(1)-like agonists on NHE3 activity. In WKY rats, fenoldopam increased G(s)alpha/NHE3 binding to the same extent in 2-wk-old (1.5-fold, n = 4) and adult (1.5-fold, n = 4) rats. In contrast, in SHRs, fenoldopam decreased the amount of G(s)alpha bound to NHE3 in 2-wk-old SHRs and had no effect in 4-wk-old and adult SHRs. These studies indicate that the decreased inhibitory effect of D(1)-like agonists on NHE3 activity in SHRs (compared with WKY rats) precedes the development of hypertension. This may be caused, in part, by a decreased interaction between G(s)alpha and NHE3 in BBM secondary to impaired D(1)-like receptor function.     

On the role of NMDA receptors in blood pressure regulation in spontaneously hypertensive rats (SHR)

Summary In the present study the binding of [³H]MK-801 to glutamatergic receptors of the NMDA type was compared in spontaneously hypertensive (SHR) and normotensive (WKY) rats in various brain structures (including nucleus tractus solitarii) by quantitative receptor autoradiography. Additionally, blood pressure changes after treatment with the NMDA antagonist MK-801 were studied in both strains. There were no differences between SHR and WKY rats either in the level of [³H]MK-801 binding or in the hypertensive reaction to MK-801.     

Effect of exercise training on aortic collagen content in spontaneously hypertensive rats (SHR)

We investigated the effects of exercise training on the amount of aortic collagen and systolic blood pressure in spontaneously hypertensive rats (SHR). Ten-week old SHR were trained either by forced treadmill running (26.8 m X min-1 -1 h X day-1, five times a week, 0% incline) or by voluntary running in revolving wheels (7,800 m X day-1 at peak) for 8 weeks. Succinate dehydrogenase (SDH) activity measured as a marker of an endurance training effect was 13% higher (P less than 0.01) in the soleus of forced-exercised animals than in that of sedentary ones. (6.56 +/- 0.17 mumol X g-1 X min-1; mean +/- SEM), whereas SDH activity in that of voluntarily-exercised group was found to be at the same level as in sedentary animals. The systolic blood pressure after training increased by 26.4 in sedentary, 21.1 in voluntarily-exercised, and 33.9 mm Hg in forced-exercised rats, when compared with the value of each group at the beginning of the training program. A significant difference was observed in the increment of blood pressure only between the voluntarily- and forced-exercised groups (P less than 0.05). The amount of aortic collagen in voluntarily-trained rats (96.5 +/- 2.0 mg X g tissue-1, 39.8 +/- 0.7 mg X 100 mg protein-1) was significantly less than that in forced-trained rats (P less than 0.05). These results suggest that voluntary, mild exercise training may be more effective in the reduction of collagen accumulation in the aorta associated with the suppression of blood pressure increase than forced, vigorous exercise training in SHR.     

alpha(2)-Adrenergic receptors in NTS facilitate baroreflex function in adult spontaneously hypertensive rats

We examined the effect of alpha(2)-adrenoreceptor blockade in the nucleus of the solitary tract (NTS) on baroreflex responses elicited by electrical stimulation of the left aortic depressor nerve (ADN) in urethane-anesthetized spontaneously hypertensive rats (SHR, n = 11) and normotensive Wistar-Kyoto rats (WKY, n = 11). ADN stimulation produced a frequency-dependent decrease in mean arterial pressure (MAP), renal sympathetic nerve activity (RSNA), and heart rate (HR). In SHR, unilateral microinjection of idazoxan into the NTS markedly reduced baroreflex control of MAP, RSNA, and HR and had a disproportionately greater influence on baroreflex control of MAP than of RSNA. In WKY, idazoxan microinjections did not significantly alter baroreflex function relative to control vehicle injections. These results suggest that baroreflex regulation of arterial pressure in SHR is highly dependent on NTS adrenergic mechanisms. The reflex regulation of sympathetic outflow to the kidney is less influenced by the altered alpha(2)-adrenoreceptor mechanisms in SHR.     

Collagen in the aortae of spontaneously hypertensive (SHR) rats

A biphasic pattern of collagen biosynthesis was found in the aortae of spontaneously hypertensive (SHR) rats; the time course of the rate of biosynthesis is similar to that described in the heart by Sen and Bumpus. In comparison to age-matched controls, collagen biosynthesis is elevated in the SHR rats, diminishes during the first fourteen weeks and rises again at the stage of established hypertension. In the period of established hypertension, the increased rate of collagen biosynthesis was associated with a pronounced rise of the collagen type I to type III ratio. On the other hand, in the pre-hypertensive stage, the proportion of collagen type III clearly exceeds the proportion of collagen type I in SHR rats.     

Effect of treatment on longevity in spontaneously hypertensive rats.

Continuous control of the pressure of spontaneously hypertensive rats at systolic levels of 90-100 mm Hg with antihypertensive agents significantly prolonged their life span. The mean survival time in control animals was 73 weeks as compared to 96 weeks in treated SHR (P less than .0005) with 11% of the latter surviving to approximately 3 yr of age. Cardiovascular lesions were limited almost exclusively to the control rats. The general health, body weight and reproductive functions of the experimental animals remained normal throughout the drug treatment. These results indicate that by preventing the rise in blood pressure and consequent cardiovascular complications the life span of the SHR may be prolonged to that of the normal albino rat.     

Effect of vasopressin and V1 receptors blockade on hypotensive action of ANP in normotensive (WKY) and spontaneously hypertensive rats.

The aim of the study was to find out whether vasopressin (AVP) modifies hypotensive and heart rate accelerating effects of atrial natriuretic peptide (ANP) in normotensive (WKY) and spontaneously hypertensive (SHR) conscious rats. The effect of i.v. administration of 1; 2 and 4 micrograms of ANP on blood pressure (MP) and heart rate (HR) was compared during i.v. infusion of 0.9% NaCl (NaCl), NaCl+AVP (1.2 ng kg-1 min-1) and NaCl+dEt2AVP (V1 receptors antagonist, 0.5 microgram kg-1 min-1). AVP increased MP in SHR and WKY and decreased HR in SHR. V1 antagonist decreased MP and increased HR only in SHR. In SHR ANP decreased MP and increased HR during NaCl, AVP and V1 antagonist infusion. In WKY these effects were observed only during AVP administration. In each experimental situation hypotension and tachycardia induced by ANP were greater in SHR than in WKY. In both strains ANP induced changes in MP and HR were enhanced during AVP in comparison to NaCl infusion. V1 antagonist did not modify effects of ANP in WKY and SHR. The results indicate that ANP abolishes hypertensive response induced by blood AVP elevation and that the basal levels of endogenous vasopressin acting through V1 receptors does not interfere with hypotensive action of ANP neither in WKY nor in SHR.     

Selective proliferation of brain kappa opiate receptors in spontaneously hypertensive rats

The binding of tritiated ligands for various opiate receptor subtypes to brain membranes prepared from spontaneously hypertensive rats and normotensive Wistar-Kyoto rats was determined. The density (Bmax) or the apparent dissociation constant (Kd) for the binding of the mu-ligand (naltrexone) and delta-ligand (Tyr-D-Ser-Gly-Phe-Leu-Thr) to brain membranes of hypertensive and normotensive rats did not differ. However, the Bmax for the binding of kappa-ligand (ethylketocyclazocine, EKC) to brain membranes after the suppression of mu and delta-sites by 100 nM each of unlabeled D-Ala2-MePhe4-Gly-ol5-enkephalin and D-Ala2-D-Leu5-enkephalin, respectively, was significantly greater in hypertensive rats compared to normotensive rats. The Kd values for the binding of 3H-EKC in the two groups did not differ. The binding of 3H-EKC in brain regions was in the order: hypothalamus greater than midbrain greater than striatum greater than cortex greater than pons + medulla. The increase in the binding of 3H-EKC in the brain of hypertensive rats compared to normotensive rats was due to increased binding in the hypothalamus and cortex. These results provide for the first time evidence of selective proliferation of kappa-opiate receptors in the brain of hypertensive rats, and suggest that brain kappa-opiate receptors may play an important role in the pathophysiology of hypertension.     

Down-regulation of endothelin receptors in the ventrolateral medulla of spontaneously hypertensive rats.

The binding of [125I] sarafotoxin 6b (SRT 6b) and [125I] endothelin-1 (ET-1) to endothelin (ET) receptors of neuronal membranes prepared from cerebral cortex and ventrolateral medulla of 8 week old spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats was determined. [125I] SRT 6b bound to the membranes of cerebral cortex and ventrolateral medulla at a single high affinity site. The binding of [125I] SRT 6b in the cerebral cortex was found to be similar in SHR and WKY rats. However, in the ventrolateral medulla [125I] SRT 6b binding was found to be significantly lower in SHR as compared to WKY rats. The decreased binding was due to decrease (48%) in the Bmax values in SHR rats as compared to WKY rats. The Kd values were similar in SHR and WKY rats. [125I] ET-1 also bound to the membranes of cerebral cortex and ventrolateral medulla at a single high affinity site. The binding of [125I] ET-1 in the cerebral cortex was found to be similar in SHR and WKY rats. However, in the ventrolateral medulla [125I] ET-1 binding was found to be significantly lower in SHR as compared to WKY rats. The decreased binding was due to 36% decrease in the Bmax values in SHR rats as compared to WKY rats. The Kd values were similar in SHR and WKY rats. It is concluded that the population of ET receptors is less in the ventrolateral medulla of SHR rats and may be contributing to the regulation of blood pressure.     

Reduced number of beta-adrenergic receptors in the myocardium of spontaneously hypertensive rats.

Inotropic response to β-adrenergic stimulation of the myocardium is decreased in hypertension. A biochemical basis for this decrease was provided by the observation that the number of β-adrenergic receptors — as reflected in specific [³H]dihydroalprenolol binding — was diminished in the myocardium of spontaneously hypertensive rats without a change in the affinity of dihydroalprenolol for the binding sites or in the capacity of isoproterenol to displace dihydroalprenolol. The decline in β-adrenergic receptor numbers is not secondary to blood pressure elevation and may be related to increased sympathetic drive in spontaneously hypertensive rats.     

Inability of cyclo (Leu-Gly) to facilitate the development of tolerance to and physical dependence on morphine in the rat

The effect of cyclo (Leu-Gly), an analog of melanotropin release inhibition factor on the development of tolerance to and physical dependence on morphine in the rat was investigated. Administration of cyclo (Leu-Gly) (1 μg/rat/day) prior to and during morphine pellet implantation failed to facilitate the development of tolerance to the analgesic and hypothermic effects of morphine. Similarly the development of dependence on morphine was not facilitated by cyclo (Leu-Gly) as evidenced by changes in body weight and body temperature observed during abrupt withdrawal of morphine. These studies do not lend support to the previous observations that cyclo (Leu-Gly) and other related peptides facilitate the development of tolerance to and physical dependence on morphine.