Measurement of psychological state changes at low dopamine transporter occupancy following a clinical dose of mazindol.

RATIONALE: The beneficial effects of psychostimulant drugs in the treatment of psychiatric disorders occur because they increase the extracellular dopamine concentration by inhibiting re-uptake of extracellular dopamine at dopamine transporters. However, the psychological effects at low dopamine transporter occupancy have not been well demonstrated. OBJECTIVES: The purpose of the study was to evaluate the psychological effects, dopamine transporter occupancy, and dopamine release induced by a single oral administration of a clinical dose of mazindol. METHODS: Ten healthy male volunteers were orally administered a placebo and a clinical dose of mazindol (1.5 mg) on separate days. The psychological effects of mazindol were assessed using a visual analogue scale to detect alterations in the state of consciousness. The amount of blockade of dopamine transporters was assessed using positron emission tomography with [18F]FE-PE2I and extracellular dopamine release was measured as the amount of change in [11C]raclopride binding. RESULTS: Following administration of a clinical dose of mazindol, the dopamine transporters were blocked by 24-25 %, and the binding potential of [11C]raclopride was reduced by 2.8-4.6 %. The differences of a score measuring derealisation and depersonalization associated with a positive basic mood were significantly correlated with the change in the [11C]raclopride binding in the limbic striatum. CONCLUSIONS: A subtle alteration in the state of consciousness was detected with a correlation to the changes in the [11C]raclopride binding, which implies that a subtle alteration in extracellular dopamine concentration in the limbic striatum by a small amount of dopamine transporter occupancy can affect the state of consciousness. TRIAL REGISTRATION HTTPS://UPLOAD.UMIN.AC.JP/CGI-OPEN-BIN/CTR_E/CTR_VIEW.CGI?RECPTNO=R000009703 : UMIN000008232.

Serum ß-cryptoxanthin and ß-carotene derived from Satsuma mandarin and brachial-ankle pulse wave velocity: The Mikkabi cohort study.

BACKGROUND AND AIMS: Findings of observational studies suggest cardioprotective effects of antioxidant vitamins and carotenoids. However, recent meta-analyses failed to show the beneficial effects of supplemental intake of antioxidants on cardiovascular disease (CVD). We aimed to assess the association between CVD risk and ß-cryptoxanthin in Japan, where Satsuma mandarin, a major source of ß-cryptoxanthin, is widely consumed. METHODS AND RESULTS: This was part of the Mikkabi cohort study. Surveys were conducted at baseline, in 2003 and 2005, and on follow-up in 2006, 2009, and 2013. We examined brachial-ankle pulse wave velocity (baPWV) with a high cut-off value set at 18.3 m s(-1). Hazard ratios (HR) and 95% confidence intervals for high baPWV were estimated using a Cox proportional hazards model with adjustment for potential confounders. A total of 635 participants with baPWV of less than 18.3 m s(-1) at baseline were included in the analysis. During the follow-up period of 57,921 person-months, 99 subjects developed high baPWV. After multivariate adjustment, the HR for high baPWV in the highest tertile compared with the lowest tertile was significantly low for ß-cryptoxanthin, ß-carotene, and total carotenoids. Serum concentrations of ß-cryptoxanthin and ß-carotene were higher in people who ate Satsuma mandarin frequently. Compared with <1/d intake of Satsuma mandarin, 3-4/d was associated with a low risk of high PWV. CONCLUSION: This study indicated that ß-cryptoxanthin and ß-carotene derived from Satsuma mandarin are candidate micronutrients for preventing arteriosclerosis development. Further longitudinal and interventional studies will be required to validate the effect on CVD.

Changes in dopamine D2-receptor binding are associated to symptom reduction after psychotherapy in social anxiety disorder.

The dopamine system has been suggested to play a role in social anxiety disorder (SAD), partly based on molecular imaging studies showing reduced levels of striatal dopaminergic markers in patients compared with control subjects. However, the dopamine system has not been examined in frontal and limbic brain regions proposed to be central in the pathophysiology of SAD. In the present study, we hypothesized that extrastriatal dopamine D2-receptor (D2-R) levels measured using positron emission tomography (PET) would predict symptom reduction after cognitive behavior therapy (CBT). Nine SAD patients were examined using high-resolution PET and the high-affinity D2-R antagonist radioligand [(11)C]FLB 457, before and after 15 weeks of CBT. Symptom levels were assessed using the anxiety subscale of Liebowitz Social Anxiety Scale (LSAS(anx)). At posttreatment, there was a statistically significant reduction of social anxiety symptoms (P<0.005). Using a repeated measures analysis of covariance, significant effects for time and time × LSAS(anx) change on D2-R-binding potential (BP(ND)) were shown (P<0.05). In a subsequent region-by-region analysis, negative correlations between change in D2-R BP(ND) and LSAS(anx) change were found for medial prefrontal cortex and hippocampus (P<0.05). This is the first study to report a direct relationship between symptom change after psychological treatment and a marker of brain neurotransmission. Using an intra-individual comparison design, the study supports a role for the dopamine system in cortical and limbic brain regions in the pathophysiology of SAD.

Dietary patterns of antioxidant vitamin and carotenoid intake associated with bone mineral density: findings from post-menopausal Japanese female subjects.

UNLABELLED: Recent studies show that antioxidants may reduce the risk of osteoporosis. This study showed the associations of bone mineral density with dietary patterns of antioxidant vitamins and carotenoids. The findings suggest the combination of vitamin C and ß-cryptoxanthin intakes might provide benefit to bone health in post-menopausal Japanese female subjects. INTRODUCTION: Recent epidemiological studies show antioxidants may reduce the risk of osteoporosis, but little is known about the dietary patterns of antioxidant vitamin and carotenoid intakes and their relation with bone mineral density (BMD). METHODS: A total of 293 post-menopausal female subjects who had received health examinations in the town of Mikkabi, Shizuoka Prefecture, Japan, participated in the study. Radial BMD was measured using dual-energy X-ray absorptiometry. Dietary intakes of antioxidant vitamins and carotenoids were assessed by using a validated food-frequency questionnaire. Dietary patterns were identified on a selected set of antioxidants through principal component factor analysis. RESULTS: Three dietary patterns were identified. The "retinol" pattern, characterized by notably high intakes of preformed retinol, zeaxanthin, and vitamin E, was positively associated with the risk for low BMD. In contrast, the "ß-cryptoxanthin" pattern, characterized by notably high intakes of ß-cryptoxanthin and vitamin C, was negatively associated with low BMD. The odds ratios for low BMD in the highest tertiles of dietary intakes of preformed retinol, vitamin C, and ß-cryptoxanthin against the lowest tertiles were 3.22 [95% confidence interval (CI), 1.38-7.51], 0.25 (CI, 0.10-0.66), and 0.40 (CI, 0.17-0.92), respectively, after adjustments for confounders. However, negative associations of vitamin C and ß-cryptoxanthin with low BMD were not significant after further adjustment for intake of ß-cryptoxanthin or vitamin C, respectively. Higher intakes of both vitamin C and ß-cryptoxanthin were significantly associated with low BMD (P < 0.05). CONCLUSIONS: The combination of vitamin C and ß-cryptoxanthin may be associated with radial BMD in post-menopausal Japanese female subjects.

Bone mineral density in post-menopausal female subjects is associated with serum antioxidant carotenoids.

UNLABELLED: High intake of fruit and vegetables may reduce the risk of osteoporosis. Carotenoids exist in abundance in these foods. This study showed the association of bone mineral density with serum carotenoids. The findings suggest that beta-cryptoxanthin and beta-carotene might provide benefits to bone health in post-menopausal female subjects. INTRODUCTION: Antioxidant carotenoids are abundant in fruit and vegetables. Recent epidemiological studies show that high intakes of fruit and vegetables may reduce the risk of osteoporosis, but little is known about the association of bone mineral density (BMD) with serum carotenoids. METHODS: A total of 699 subjects (222 males and 477 females) who had received health examinations in the town of Mikkabi, Shizuoka Prefecture, Japan, participated in the study. Radial BMD was measured using dual-energy X-ray absorptiometry (DXA). The associations of serum carotenoid concentrations with the radial BMD were evaluated cross-sectionally. RESULTS: In male and pre-menopausal female subjects, the six serum carotenoids were not associated with the radial BMD. On the other hand, in post-menopausal female subjects, serum beta-cryptoxanthin and beta-carotene were weakly but positively correlated with the radial BMD. After adjustment for confounders, the odds ratio (OR) for the lowest quartile of BMD in the high groups (Q2-Q4) of serum beta-cryptoxanthin against the lowest quartile (Q1) was 0.45 (95% confidence interval: 0.22-0.95) in post-menopausal female subjects. However, this association was not significant after further adjusting for intakes of minerals and vitamins. CONCLUSIONS: Antioxidant carotenoids, especially beta-cryptoxanthin, significantly but partly associate with the radial BMD in post-menopausal female subjects.

Cloning and expression of CitPDS1, a gene encoding phytoene desaturase in citrus.

A cDNA clone encoding phytoene desaturase (PDS) was isolated from citrus (Citrus unshiu Marc.). The transcript of the isolated PDS (CitPDS1) was not detected by conventional RNA gel-blot analysis; instead, it was detected by a sensitive reverse transcription-PCR (RT-PCR). The CitPDS1 transcript in the juice sacs/segment epidermis (edible part) was at a low level in the young fruit, and it increased toward maturation like citrus phytoene synthase (CitPSY1). In the peel, in contrast to CitPSY1, the transcript of which was induced toward maturation, the level of the CitPDS1 transcript remained constant after an increase in July, indicating non-coordinate regulation of CitPDS1 and CitPSY1 in the peel.

3',5'-Di-C-beta-glucopyranosylphloretin, a flavonoid characteristic of the genus Fortunella.

Dihydrochalcone derivative, 3',5'-di-C-beta-glucopyranosylphloretin (1), is present in the genus Fortunella, (F. crassifolia, F. japonica, F. margarita, F. polyandra and F. hindsii). These species accumulate a large quantity of 1 in their fruits (peel, 6.5-15.2 mg/g in dry wt; juice sac, 1.5-10.5 mg/g) and in their leaves (21.3-60.2 mg/g). Twenty-seven Tanaka's Citrus species examined lack 1, but C. madurensis and C. halimii contain 1 in large quantities in their peels (25.1 and 33.6 mg/g) and juice sacs (4.1 and 4.2 mg/g). Poncirus species do not contain 1. Fortunella-citrus hybrids, the Orangequat [C. unshiuxF. crassifolia], the Thomasville citrangequat [Fortunella sp.x(C. sinensisxPoncirus trifoliata)], and seven hybrid progenies [F. margaritaxC. junos], contain large amounts of 1 in their peels (17.0-7.9 mg/g) and juice sacs (2.0-9.9 mg/g). These facts suggest that accumulation of 1 is a generic trait of the genus Fortunella and that the inheritance of the trait among the intergeneric hybrids is controlled by a dominant allele. Thus C. madurensis and C. halimii are thought to originate from natural hybrids between the genera Citrus and Fortunella. Phloridzin, which has the same aglycon as 1, was not detected in the citrus plants examined.

Comparison of parametric FBP and OS-EM reconstruction algorithm images for PET dynamic study.

An ordered subsets expectation maximization (OS-EM) algorithm is used for image reconstruction to suppress image noise and to make non-negative value images. We have applied OS-EM to a digital brain phantom and to human brain 18F-FDG PET kinetic studies to generate parametric images. A 45 min dynamic scan was performed starting injection of FDG with a 2D PET scanner. The images were reconstructed with OS-EM (6 iterations, 16 subsets) and with filtered backprojection (FBP), and K1, k2 and k3 images were created by the Marquardt non-linear least squares method based on the 3-parameter kinetic model. Although the OS-EM activity images correlated fairly well with those obtained by FBP, the pixel correlations were poor for the k2 and k3 parametric images, but the plots were scattered along the line of identity and the mean values for K1, k2 and k3 obtained by OS-EM were almost equal to those by FBP. The kinetic fitting error for OS-EM was no smaller than that for FBP. The results suggest that OS-EM is not necessarily superior to FBP for creating parametric images.

Cervical cancer in young Japanese women.

This study was performed to determine whether the incidence of cervical cancer in women aged 35 or younger has changed over the last 10 years and to examine the clinical characteristics of the cases. The incidence of cervical cancer in women aged 35 or younger were significantly greater in 1987-1991 than 1992-1996 (p = 0.001). Most new cases were detected by routine cytological screening.

Wound-induced ethylene synthesis and expression and formation of 1-aminocyclopropane-1-carboxylate (ACC) synthase, ACC oxidase, phenylalanine ammonia-lyase, and peroxidase in wounded mesocarp tissue of Cucurbita maxima.

1-Aminocyclopropane-1-carboxylate (ACC) synthase was rapidly induced in mesocarp tissue of Cucurbita maxima after wounding in the cut surface layer in 1 mm thickness (ca. 9 cells) (first layer) in both the enzyme activity and the levels of transcript. This led to a rapid accumulation of ACC and hence ethylene production. In the inside tissue (1-2 mm) (second layer), no significant induction of ACC synthase was observed, which resulted in a low level of ACC, although ethylene was evolved at a much lower rate than the first one. In contrast to ACC synthase, ACC oxidase was induced markedly in both the first and second layers and the development of its activity and the levels of mRNA remained high until later stages. It was considered that wound ethylene was closely associated with the development of ACC oxidase, since 2,5-norbornadiene (NBD), an inhibitor of ethylene action, substantially suppressed it. Phenylalanine ammonia-lyase (PAL) greatly increased in activity after wounding similarly to that of ACC synthase, in which increase in PAL activity occurred predominantly in the first layer. Induction of peroxidase activity after wounding had a close correlation in profile with that of ACC oxidase in that marked increases in the activity were observed in both the first and second layers and were strongly suppressed by NBD application. Four peroxidase isozymes were found by PAGE, among which a fraction was newly detected after wounding.

Evaluation of auraptene content in citrus fruits and their products.

Auraptene quantities in Tanaka's 77 Citrus species (including 14 varieties and cultivars), 5 Fortunella species, one Poncirus species, 27 hybrids between Citrus species, and 51 intergeneric hybrids between Citrus and Poncirus have been evaluated. The genus Citrus has been divided into eight groups. Auraptene is found in all of the species of Cephalocitrus group, a part of the species of Aurantium group, and most of the species of Osmocitrus group. The Citrus species contain a small amount of auraptene in the juice sacs compared with in the peels except for Henka mikan (C. pseudo-aurantium), Ichang lemon (C. wilsonii), and a Hassaku (C. hassaku)-pummelo hybrid (Okitsu No. 39), which contain large quantities of auraptene in their juice sacs (0.23, 0.52, and 0.14 mg/g, respectively). The Hong Kong wild kumquat (F. hindusii) alone contains auraptene in Fortunella species. All of the Citrus-trifoliate orange (P. trifoliata) hybrids as well as the trifoliate orange contain a large quantity of auraptene in both the peel (16.57-0.51 mg/g) and the juice sac (10.32-0.15 mg/g). These hybrids are almost inedible. The Iyo (C. iyo)-trifoliate orange hybrid (IyP269) is edible and contains auraptene in the peel (1.49 mg/g) and in the juice sac (1.73 mg/g). Citrus fruit products, for example, brand-named grapefruit juice and marmalade, retain about 0. 1 mg and 0.3 mg/100 g of auraptene, respectively.

cDNA sequence and expression of a cold-responsive gene in Citrus unshiu.

A cDNA clone encoding a protein (CuCOR19), the sequence of which is similar to Poncirus COR19, of the dehydrin family was isolated from the epicarp of Citrus unshiu. The molecular mass of the predicted protein was 18,980 daltons. CuCOR19 was highly hydrophilic and contained three repeating elements including Lys-rich motifs. The gene expression in leaves increased by cold stress.

Characterization of a cDNA homologous to carotenoid-associated protein in citrus fruits.

A cDNA (CitPAP) homologous to a gene encoding for Cucumis sativus carotenoid-associated protein (CHRC) has been isolated from satsuma mandarin (Citrus unshiu Marc.). Unlike ChrC whose expression was limited only in mature fruits (containing chromoplasts), CitPAP transcripts were detected in all the tissues examined including fruits, flowers and leaves. In this respect, CitPAP was rather close to a gene encoding for pepper plastid-lipid-associated protein (PAP), which exhibits ubiquitous expression in bell pepper organs containing chloroplasts or chromoplasts. CitPAP, however, differed from PAP in the magnitude and pattern of RNA accumulation. These results might indicate a novel function of CitPAP.

[Creation of a dynamic digital phantom and its application to a kinetic analysis].

A dynamic digital brain phantom was created from the MRI to evaluate visually the relationship between the noise and the error in the parameter estimates in the PET kinetic analysis. This phantom incorporates the noise level depending on administration dose, camera efficiency and the data acquisition schedule. We simulated a serial dynamic scan with 18F-FDG or 11C-flumazenil, assuming 2-tissue 3-parameter model and 1-tissue 2-parameter model, respectively, and the sampling schedule was determined according to the clinical examination. The noise in the tissue time activity curve in FDG had a peak in the first minute and decreased thereafter, whereas the noise increased gradually in the flumazenil study after the initial peak due to radioactivity decay. We examined the relationship between the noise level and the error in the parameter estimates. Both mean absolute differences between true and estimated values and standard deviation became large, and the quality of the parametric images became poor with increasing noise level. This simulation was compared with human tissue time activity curves and parametric images, which were obtained with 100 MBq administration dose in FDG study and 430 MBq in flumazenil study. We inferred that the noise level in the human study was 10-20% in FDG, and 20-40% in flumazenil, and the error in the estimated parameter of K-complex in FDG study was about 20%, that of DV in flumazenil study was 2%.

[A case of adenoid cystic carcinoma presenting with stridor and which was treated by reversed gamma type stent placement].

A 66-year-old man was admitted to Saiseikai Suita Hospital with stridor and dyspnea. The flow-volume curve showed central airway obstruction pattern. Although a P-A chest roentogenogram was normal, chest tomogram demonstrated a tumor shadow in the lower portion of the trachea. Chest CT examination demonstrated a tumor arising from the posterior wall of the lower portion of the trachea and stenosis with wall thickening extending from the lower portion of the trachea to both main bronchi. Bronchofiberscopic examination revealed polypoid tumors obstructing the lower portion of the tracheal lumen, and biopsy was subsequently performed. On the night immediately after the bronchofiberscopic examination, the patent suddenly choked, and emergency intubation was performed. Adenoid cystic carcinoma was diagnosed on biopsy and we decided to place a spiral Z-stent. A reversed gamma-type stent placement was used. The first spiral Z-stent extended from the trachea to the right main bronchus and the second spiral Z-stent was placed in the left main bronchus through the first stent. Following the spiral Z-stent placement radiotherapy was performed. Spiral Z-stent placement resulted in dilation of the stenotic trachea and both main bronchi.

Peroxisome targeting signal type 1 (PTS1) receptor is involved in import of both PTS1 and PTS2: studies with PEX5-defective CHO cell mutants.

To investigate the mechanisms of peroxisome assembly and the molecular basis of peroxisome assembly disorders, we isolated and characterized a peroxisome-deficient CHO cell mutant, ZP139, which was found to belong to human complementation group II, the same group as that of our earlier mutant, ZP105. These mutants had a phenotypic deficiency in the import of peroxisomal targeting signal type 1 (PTS1) proteins. Amino-terminal extension signal (PTS2)-mediated transport, including that of 3-ketoacyl coenzyme A thiolase, was also defective in ZP105 but not in ZP139. PEX5 cDNA, encoding the PTS1 receptor (PTS1R), was isolated from wild-type CHO-K1 cells. PTS1R's deduced primary sequence comprised 595 amino acids, 7 amino acids less than the human homolog, and contained seven tetratricopeptide repeat (TPR) motifs at the C-terminal region. Chinese hamster PTS1R showed 94, 28, and 24% amino acid identity with PTS1Rs from humans, Pichia pastoris, and Saccharomyces cerevisiae, respectively. A PTS1R isoform (PTS1RL) with 632 amino acid residues was identified in CHO cells; for PTS1R, 37 amino acids were inserted between residues at positions 215 and 216 of a shorter isoform (PTS1RS). Southern blot analysis of CHO cell genomic DNA suggested that these two isoforms are derived from a single gene. Both types of PEX5 complemented impaired import of PTS1 in mutants ZP105 and ZP139. PTS2 import in ZP105 was rescued only by PTS1RL. This finding strongly suggests that PTS1RL is also involved in the transport of PTS2. Mutations in PEX5 were determined by reverse transcription-PCR: a G-to-A transition resulted in one amino acid substitution: Gly298Glu of PTS1RS (G335E of PTS1RL) in ZP105 and Gly485Glu of PTS1RS (G522E of PTS1RL) in ZP139. Both mutations were in the TPR domains (TPR1 and TPR6), suggesting the functional consequence of these domains in protein translocation. The implications of these mutations are discussed.

[Cytokine producing ability of peripheral blood mononuclear cells in the clinical course of pulmonary tuberculosis].

Interferon-gamma (IFN-gamma) and interleukin-10 (IL-10)-producing ability of peripheral blood plastic-dish adherent cells and non-adherent cells obtained from patients with active pulmonary tuberculosis (N = 17) and healthy controls (N = 14) upon stimulation with purified protein derivatives (PPD) were assessed. Adherent cells and non-adherent cells were obtained two times from each patient with active pulmonary tuberculosis without any underlying diseases, on admission before the initiation of administering anti-tuberculous drugs and 2 months later from the negative conversion of Mycobacterium tuberculosis in sputum culture. ELISA was performed to measure IFN-gamma and IL-10 levels in culture media of adherent cells and non-adherent cells stimulated with PPD. IFN-gamma levels produced by non-adherent cells on admission were significantly higher than that of healthy controls (p < 0.001). Elevated IFN-gamma levels on admission was reduced after treatment for tuberculosis (p < 0.03), but still remained higher than that in healthy controls. IL-10 levels of non-adherent cells of patients were lower than those of healthy controls, although the difference was not significant. IL-10 levels produced by non-adherent cells on admission correlated with the time needed for negative conversion of bacilli in sputum culture (p < 0.05). IL-10 level produced by adherent cells from nutritionally normal patients were significantly higher than that of healthy controls (p < 0.05), and elevated IL-10 level was significantly reduced after therapy (p < 0.05). In the normonourished patients, the time needed for negative conversion of the bacilli in sputum culture of patients kept higher level of IL-10 of non-adherent cells (N = 5) was significantly longer than that of patients reduced IL-10 level after therapy. These results suggest that IL-10 produced by monocytes may diminish the TH1 responses of patients with pulmonary tuberculosis.

Familial macrothrombocytopenia with unusually elongated mitochondria.

We report a familial case of macrothrombocytopenia without inclusion bodies in polymorphonuclear cells or any congenital abnormalities. The results of the hemostatic and platelet function tests were all normal except for the platelet retention rate. The number of megakaryocytes increased slightly and some were relatively small. Electron microscopic studies revealed a unique morphological abnormality of the platelets' mitochondria.

[Effects of terguride, an ergot alkaloid derivative, on the central nervous system: biochemical and behavioral studies].

Effects of terguride, a 9,10-dihydrogenated derivative of lisuride, on the central nervous system were investigated in rodents in comparison with those of lisuride. In vitro binding studies in rat brains showed that terguride, similar to lisuride, had a high affinity for D2-, 5-HT1A-, 5-HT2-, alpha 1- and alpha 2-receptors. Terguride, as does lisuride, induced hypomotility and yawning at low doses in rats, suggesting its presynaptic D2-agonist action. Terguride, unlike the postsynaptic D2-agonist lisuride, induced neither hypermotility nor stereotypy in rats and guinea pigs, but suppressed the hypermotility and stereotypy induced by apomorphine. Terguride suppressed haloperidol-induced catalepsy in rats and induced contralateral rotations in unilaterally 6-OHDA-lesioned rats, as does lisuride. These effects may be due to the postsynaptic D2 partial agonist action. Terguride, unlike lisuride, neither induced the serotonin syndrome nor generalized to the discriminative stimuli of the 5-HT1A- agonist 8-OH-DPAT in rats. Terguride did not induce head twitch in mice. Terguride blocked noradrenaline-induced lethality and clonidine-induced hypothermia at high doses in mice. Repeated administration of terguride did not affect the behavioral actions in rats. Thus, the effects of terguride on the central nervous system seems to be produced by mediation of the agonist and partial agonist actions at presynaptic and postsynaptic D2- receptors, respectively.

Effects of prolonged application of isoprenaline on intracellular free magnesium concentration in isolated heart of rat.

1. The effect of prolonged application of isoprenaline on intracellular free-Mg2+ concentration ([Mg2+]i) was examined by use of 31P-nuclear magnetic resonance (31P-n.m.r.) in rat isolated hearts. Left ventricular pressure (LVP) was simultaneously measured. 2. [Mg2+]i was estimated from the separation of the alpha- and beta-ATP peaks, using the dissociation constant of MgATP 38 microM (established previously). In normal (phosphate-free, Krebs-Henseleit) solution, [Mg2+]i was approximately 0.4 mM. 3. When isoprenaline was applied for 100 min, a transient increase in [Mg2+]i was observed during the initial 25 min, whilst concentrations of ATP ([ATP]) and phosphocreatine ([PCr]) decreased and [Pi] correspondingly increased. During the subsequent 75 min of isoprenaline application, [Mg2+]i decreased below its resting levels. Washout of isoprenaline restored [Mg2+]i and [PCr], but [ATP] remained low. These changes elicited by isoprenaline were not observed in the presence of propranolol, a typical alpha-adrenoceptor blocker. 4. Isoprenaline increased both LVP and heart rate. The increased LVP and heart rate slowly returned to lower values during prolonged application of isoprenaline, but remained higher than those before application. 5. The transient rise in [Mg2+]i elicited by isoprenaline could be attributed to the decrease in [ATP] resulting in a release of Mg2+. The subsequent decrease in [Mg2+]i during the prolonged applications suggests that beta-adrenoceptor stimulation itself facilitates Mg(2+)-extruding mechanism(s).