Dietary potassium and magnesium supplementation in cyclosporine-induced hypertension and nephrotoxicity.

BACKGROUND: Cyclosporine A (CsA)-induced hypertension and nephrotoxicity are aggravated by high sodium intake. Accumulating evidence suggests that potassium and magnesium supplementation could protect against the detrimental effects of dietary salt. In the present study, we tested the hypothesis of whether concurrent supplementation with potassium and magnesium could protect against the development of CsA-induced hypertension and nephrotoxicity more effectively than supplementation with one mineral alone. METHODS: Eight-week-old spontaneously hypertensive rats (SHRs) were divided into four groups (N = 10 in each group): (1) CsA group (5 mg/kg subcutaneously) receiving high-sodium diet (Na 2.6%, K 0.8%, Mg 0.2% wt/wt); (2) CsA group receiving a high-sodium, high-potassium diet (Na 2.6%, K 2.4%, Mg 0.2%); (3) CsA group receiving high-sodium, high-magnesium diet (Na 2.6%, K 0.8%, Mg 0.6%); and (4) CsA group receiving high-sodium, high-potassium, high-magnesium diet (Na 2.6%, K 2.4%, Mg 0.6%). RESULTS: CsA induced severe hypertension and deteriorated renal functions in SHRs on high-sodium diet. Histologically, the kidneys showed severe thickening of the media of the afferent artery with fibrinoid necrosis. Potassium supplementation lowered blood pressure (198 +/- 5 vs. 212 +/- 2 mm Hg, P < 0.05) and partially prevented the development of proteinuria (-25%, P < 0.05). Magnesium supplementation decreased blood pressure to the same extent but improved renal functions more effectively than potassium. The greatest protection against CsA toxicity was achieved when dietary potassium and magnesium supplementations were combined. Urinary N-acetyl-beta-D-glucosaminidase (NAG) excretion, a marker for renal proximal tubular damage, increased progressively in CsA-treated SHRs on the high-sodium diet. Neither potassium nor magnesium influenced urinary NAG excretion. We also estimated the activity of the renal dopaminergic system by measuring 24-hour urinary dopamine excretion rates. CsA suppressed the renal dopaminergic system during high-sodium diet. Magnesium supplementation, alone and in combination with potassium, protected against the development of renal dopaminergic deficiency in CsA-treated SHRs on high-sodium diet. Magnesium supplementation increased plasma-free ionized magnesium (iMg) and bone magnesium by 50 and 16%, respectively. CONCLUSIONS: Our findings indicate that both potassium and magnesium supplementations showed beneficial effects against CsA-induced hypertension and nephrotoxicity. The protective effect of magnesium clearly exceeded that of potassium. The greatest protection against CsA toxicity was achieved when potassium and magnesium were combined. We also provide evidence that the development of CsA-induced glomerular, tubular, and vascular lesions are associated with renal dopaminergic deficiency.

Optimization of the hydrolysis of conjugated L-DOPA, dopamine and dihydroxyphenylacetic acid in human urine for assay by high-performance liquid chromatography with electrochemical detection.

Conjugates of the catechol compounds, L-dihydroxyphenylalanine (L-DOPA), dopamine and dihydroxyphenylacetic acid in human urine were analysed using the isocratic ion-pair reversed-phase HPLC method with electrochemical detection. Acid hydrolysis, using 4 mol/l HCl for 60 min, was more effective than treatment with sulphatase for the generation of free catechols. Free (non-conjugated) catechols already present, as well as those produced by either of the hydrolysis procedures, were adsorbed onto aluminium oxide and extracted in acid solution. The repeatability of the technique for within and between-batch urine analysis was less than 2% and 8%, respectively. Free urinary dopamine (and dihydroxyphenylacetic acid) concentrations were much higher in the urine of patients treated with L-DOPA for Parkinson's disease than in healthy volunteers. At high dopamine (and dihydroxyphenylacetic acid) levels the conjugation capacity was apparently exceeded, since the overall percent conjugation of L-DOPA, dopamine and dihydroxyphenylacetic acid was decreased "concentration dependently" where the concentrations of free catechols were increased. Both in the control group and L-DOPA-treated groups, enzymatic hydrolysis was much less effective than acid hydrolysis in generating free catechols. This indicated that there were other, non-sulphated conjugates in the urine, accounting for between 66 and 100% of total conjugates.

Morphine withdrawal alters anterior pituitary hormone secretion, brain endopeptidase activity and brain monoamine metabolism in the rat.

Rats were made tolerant to morphine by a 5-day regimen with increasing doses. The time course of changes in serum anterior pituitary hormone levels, brain endo- and exopeptidase activity, levels of brain biogenic amines and body weight were studied during abrupt morphine withdrawal. Cold stimulated secretion of thyrotropin and the secretion of growth hormone were both decreased whereas that of prolactin was increased. In the hypothalamus both prolyl endopeptidase and dipeptidyl peptidase IV activities were concomitantly increased. The hypothalamic 5 hydroxyindole acetic acid levels were also increased. Changes in hormone secretion, peptidase activity and monoamine turnover had returned to baseline levels by 92 hr. Our results indicate that morphine withdrawal and the associated stress produce alterations in anterior pituitary thyrotropin and growth hormone secretion. Concomitant increases in hypothalamic prolyl endopeptidase and dipeptidyl peptidase activities may contribute to these changes.

Variation in tolerance to the antinociceptive, hormonal and thermal effects of morphine after a 5-day pre-treatment of male rats with increasing doses of morphine.

The manifestation of tolerance to the effects of morphine on nociception and the secretion of anterior pituitary hormones, and the correlation of hormonal effects to changes in body temperature and to hypothalamic monoamines were studied in male rats. Morphine (three times a day in increasing doses) or saline (control) were administered intraperitoneally during a 5-day treatment and either saline or morphine was administered as an acute challenge 92 h later. The influence of the thermal environment on the effect of morphine on the body temperature was also studied. The 5-day morphine regimen was sufficient for the development of tolerance to the antinociceptive effect of morphine. After a 92-h lag-time, the tolerance was still complete. Tolerance to the depressant effect of morphine (10-25 mg/kg) on cold-stimulated TSH secretion was seen at 2 h, but was only barely detectable at 1 h, after the injection of a challenge dose. On the other hand, a tolerance to the stimulatory effect of morphine on prolactin secretion was already seen 1 h after the acute dose of morphine. Tolerance to the hypothermic effect of morphine (25 mg/kg) was evident in rats kept at +4 degrees C after the challenge dose. On the contrary, no tolerance to the hyperthermic effect of morphine (15 or 25 mg/kg) was observed in rats kept at +30 degrees C. However, the hyperthermia was reversed when these rats were moved to +4 degrees C for 30 min, irrespective of whether they were morphine pretreated or not. Thus the removal of the hyperthermic stimulus decreased the core temperature of all rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Development of tolerance to the hormonal effects of morphine without changes in the aminergic functions in the brain of the rat.

The effect of morphine on cold-stimulated secretion of TSH and prolactin was studied in male rats, both in acute studies and after the chronic administration of morphine for 14 days (twice a day with increasing doses). The duration of the stimulatory effect of a single dose of morphine on secretion of prolactin was shorter (less than 2 hr) than its inhibitory effect on cold-stimulated secretion of TSH (over 2 hr). In the rats pretreated with morphine, a tolerance to the depressant effect of TSH of the challenge dose of morphine was seen at 2 hr but not at 1 hr after the injection. In contrast, a tolerance to the stimulatory effect of morphine on prolactin was seen at 1 hr after the acute dose of morphine. The minor alterations of the hypothalamic amine neurotransmitters and their metabolites did not correlate with the hormonal responses or to the development of tolerance.

Effect of nitecapone and clorgyline, given intracerebro-ventricularly on L-dopa metabolism in the rat brain.

A new COMT inhibitor, nitecapone (OR-462) or clorgyline, a MAO-A inhibitor, was infused into the 3rd brain ventricle (i.c.v.) of conscious male rats. None of the enzyme inhibitors given alone alter hypothalamic or striatal levels of L-dopa, dopamine or their metabolites. Most of the rats were pretreated with levodopa/carbidopa (LD/CD, 15/30 mg kg-1 intraperitoneally). Now, the action of nitecapone is localized in the hypothalamus since homovanillic acid (HVA) is decreased there, not in the striatum. The levels of 3-O-methyldopa (3-OMD) are not changed in either brain region, suggesting a lack of the peripheral leakage of nitecapone. Clorgyline (3 and 10 micrograms rat-1) elevates hypothalamic and dopamine levels. Nitecapone and clorgyline decrease prolactin (PRL) levels below those reduced by LD/CD treatment.

Different in vivo properties of three new inhibitors of catechol O-methyltransferase in the rat.

1. We compared three new catechol O-methyltransferase (COMT) inhibitors (OR-611, Ro 40-7592 and CGP 28014; 10 and 30 mg kg-1, i.p.) in male rats given levodopa (L-DOPA, 50 mg kg-1, i.p.) and carbidopa ((-)-L-alpha-methyl dopa, 50 mg kg-1, i.p.). In some studies pretreatment with pargyline (80 mg kg-1, i.p.) was used to block the function of monoamine oxidase (MAO). 2. Decreases of hypothalamic and striatal 3-O-methyl-dopa (3-OMD) levels were used as measures of the inhibition of peripheral COMT. The inhibition of brain COMT activity was estimated by decreases of hypothalamic and striatal homovanillic acid (HVA) and 3-methoxytyramine (3-MT; after pargyline) levels. 3. The three COMT inhibitors studied had different individual characteristics. OR-611 was primarily a peripherally acting COMT inhibitor, decreasing 3-OMD levels in the striatum (to 31-52%) and in the hypothalamus (to 16-27%) both in the control and pargyline-treated animals at 1 and 3 h. It did not have any effect on brain HVA and 3-MT. 3. Ro 40-7592 was a broad spectrum COMT inhibitor decreasing striatal and hypothalamic 3-OMD (always to less than 30%), HVA (to less than 50%) and 3-MT levels (to less than 23%) significantly both at 1 and 3 h. It was more potent than OR-611. 4. CGP 28014 functioned as a weak COMT inhibitor in the periphery inhibiting 3-OMD formation only at 3 h. In contrast, it was fairly potent in decreasing the brain HVA and 3-MT levels at 1 h (to 37-22% and 42-35% in the striatum, and to 57-33% and 64-35% in the hypothalamus, respectively) but not at 3 h. Since CGP 28014, unlike OR-611 and Ro 40-7592, did not generally increase the brain DOPA, dopamine or DOPAC levels, it was not a typical COMT inhibitor.

Effects of systemic carbidopa on dopamine synthesis in rat hypothalamus and striatum.

Significant concentrations of carbidopa (CD) were found in rat hypothalamus, striatum, and in striatal microdialysis efflux after intraperitoneal administration of the drug. Efflux levels peaked one hour after administration of 100 mg/kg at 0.37 micrograms/ml, or about 2% of serum levels. Concurrent CD levels in hypothalamus and striatum were about 2.5% and 1.5%, respectively, of corresponding serum levels. Levels of dopamine and its principal metabolites in striatal efflux were unaffected. The removal of the brain blood by saline perfusion decreased the striatal and hypothalamic CD concentrations only by 33% and 16%, respectively. In other rats receiving both CD and levodopa (LD), brain L-dopa, dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) levels after one hour tended to be proportionate to LD dose. When the LD dose remained constant, increasing the CD dose dose-dependently enhanced L-dopa levels in the hypothalamus and striatum. However dopamine levels did not increase but, in contrast, decreased dose-dependently (although significantly only in the hypothalamus). CD also caused dose-dependent decrease in striatal 3-O-methyldopa (3-OMD) and in striatal and hypothalamic homovanillic acid (HVA), when the LD dose was 50 mg/kg. We conclude that, at doses exceeding 50 mg/kg, sufficient quantities of CD enter the brain to inhibit dopamine formation, especially in the hypothalamus. Moreover, high doses of LD/CD, both of which are themselves catechols, can inhibit the O-methylation of brain catecholamines formed from the LD.

Effect of high single doses of levodopa and carbidopa on brain dopamine and its metabolites: modulation by selective inhibitors of monoamine oxidase and/or catechol-O-methyltransferase in the male rat.

The upper limits of striatal and hypothalamic dopamine formation and metabolism in the rat were defined after acute levodopa/carbidopa (100/100 mg/kg) in combination with MAO (clorgyline; 32 mg/kg or pargyline; 100 mg/kg) and/or COMT inhibitors (OR-462, OR-611, Ro 41-0960, 30 mg/kg). Striatal and hypothalamic dopa and 3-OMD levels increased several hundred times after levodopa/carbidopa treatment alone. Dopamine, DOPAC, HVA and 3-MT levels elevated also but noradrenaline and 5-HT did not. Clorgyline further increased 3-OMD, dopamine and 3-MT concentrations while DOPAC and HVA levels decreased. These changes were even more pronounced after pargyline. In the striatum, all COMT inhibitors (with levodopa/carbidopa) blocked 3-OMD formation but elevated neither dopamine nor DOPAC levels. OR-462 increased dopa levels. Only Ro 41-0960, the brain penetrating compound, blunted HVA levels. All three COMT inhibitors decreased high 3-OMD levels evoked by MAO inhibitors (+ levodopa/carbidopa). In pargyline-treated rats, COMT inhibitors did not alter dopamine, DOPAC or HVA levels but all of them decreased significantly 3-MT levels, particularly Ro 41-0960. Striatal dopamine levels increased maximally 6 times compared to those in the saline-treated controls. In the hypothalamus, COMT inhibitors decreased 3-OMD levels to 1/5-1/30 of those after levodopa/carbidopa alone. COMT inhibitors suppressed 3-OMD formation also in clorgyline and pargyline (+ levodopa/carbidopa) treated rats. After clorgyline, OR-611 and Ro 41-0960 increased high dopamine levels but only Ro 41-0960 suppressed HVA and 3-MT levels. None of the COMT inhibitors changed the high dopamine and low DOPAC levels after pargyline.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of hypothalamic paraventricular nucleus lesion on the cold-stimulated TSH responses to 5-HT in male rats.

The effects of lesion of the hypothalamic paraventricular nuclei (PVNx), the main thyrotrophic area, on the cold-stimulated thyrotropin (TSH) responses to intracerebroventricular (i.c.v.) 5-HT were studied in male rats. PVNx significantly attentuated the cold-stimulated TSH levels, but significantly affected neither hypothalamic thyrotropin-releasing hormone nor somatostatin content. Serum T3 levels were significantly decreased 8 days after PVNx. Irrespective of the lesion (sham or PVNx), 5-HT infusion (9 micrograms per rat) into the posterior third ventricle attenuated markedly the cold-stimulated TSH levels, whereas infusion into the anterior third ventricle did not. Bilateral 5-HT infusions (2 micrograms per side) into the hypothalamic dorsomedial nuclei significantly decreased serum TSH, but bilateral infusions into the posterior hypothalamic nuclei were without effect. Sham-lesion and PVNx decreased serum prolactin levels without affecting the stimulation of prolactin secretion by i.c.v. 5-HT. These results suggest that the inhibitory effect of i.c.v. 5-HT on TSH secretion and its stimulatory action on prolactin secretion are only partially dependent on the PVN.

Effect of acute levodopa on brain catecholamines after selective MAO and COMT inhibition in male rats.

Interactions between a selective catechol-O-methyltransferase (COMT) inhibitor OR-462 and a monoamine oxidase (MAO)-A inhibitor clorgyline were studied measuring concentrations of L-dopa, dopamine and their metabolites in the rat hypothalamus and striatum after administration of levodopa/carbidopa (15/30 mg/kg i.p.). Part of the experiments were performed in rats pretreated with 6-OH-dopamine (6-OHDA) intracerebroventricularly (i.c.v.) to determine whether changes in dopamine metabolism occurred inside or outside catecholaminergic neurons. OR-462 was an effective COMT inhibitor at the doses 3 and 30 mg/kg i.p. Inhibition of 3-O-methyldopa (3-OMD) formation from L-dopa was reflected in the hypothalamus (45-81% decrease) and striatum (87-88% decrease), since 3-OMD penetrates the blood-brain barrier. Homovanillic acid (HVA) was decreased only in the striatum at 30 mg/kg of OR-462. Clorgyline (8 and 32 mg/kg i.p.) decreased 3,4-dihydroxyphenylacetic acid (DOPAC) formation in the hypothalamus and striatum by 61-91%. When given together, OR-462 and clorgyline elevated hypothalamic dopamine levels 3.2-4.6-fold, but striatal dopamine only 1.3-1.9-fold. The formation of 3-OMD and DOPAC remained suppressed and even brain HVA levels were decreased by 51-97%. 6-OHDA treatment decreased striatal and hypothalamic dopamine by 50% and noradrenaline by 75%. In these animals levodopa/carbidopa increased brain L-dopa 2.4-4-fold, those of 3-OMD 1.2-1.7-fold compared to intact animals, but the synthesis and metabolism of dopamine and the effects of COMT and MAO inhibitors were not significantly changed. Levodopa/carbidopa treatment decreased significantly prolactin and thyrotropin levels in serum but none of the additional treatments changed this action.