Salbutamol-induced electrophysiological changes show no correlation with electrophysiological changes during hyperinsulinaemic-hypoglycaemic clamp in young people with Type 1 diabetes.

AIMS: Hypoglycaemia causes QT-interval prolongation and appears pro-arrhythmogenic. Salbutamol, a ß2 -adrenoreceptor agonist also causes QT-interval prolongation. We hypothesized that the magnitude of electrophysiological changes induced by salbutamol and hypoglycaemia might relate to each other and that salbutamol could be used as a non-invasive screening tool for predicting an individual's electrophysiological response to hypoglycaemia. METHODS: Eighteen individuals with Type 1 diabetes were administered 2.5 mg of nebulized salbutamol. Participants then underwent a hyperinsulinaemic-hypoglycaemic clamp (2.5 mmol/l for 1 h). During both experiments, heart rate and serum potassium (and catecholamines during the clamp) were measured and a high-resolution electrocardiogram (ECG) was recorded at pre-set time points. Cardiac repolarization was measured by QT-interval duration adjusted for heart rate (QTc ), T-wave amplitude (Tamp ), T-peak to T-end interval duration (Tp Tend ) and T-wave area symmetry (Tsym ). The maximum changes vs. baseline in both experiments were assessed for their linear dependence. RESULTS: Salbutamol administration caused QTc and Tp Tend prolongation and a decrease in Tamp and Tsym . Hypoglycaemia caused increased plasma catecholamines, hypokalaemia, QTc and Tp Tend prolongation, and a decrease in Tamp and Tsym . No significant correlations were found between maximum changes in QTc [r = 0.15, 95% confidence interval (95% CI) -0.341 to 0.576; P = 0.553), Tp Tend (r = 0.075, 95% CI -0.406 to 0.524; P = 0.767), Tsym (r = 0.355, 95% CI -0.132 to 0.706; P = 0.149) or Tamp (r = 0.148, 95% CI -0.347 to 0.572; P = 0.558) in either experiment. CONCLUSIONS: Both hypoglycaemia and salbutamol caused pro-arrhythmogenic electrophysiological changes in people with Type 1 diabetes but were not related in any given individual. Salbutamol does not appear useful in assessing an individual's electrophysiological response to hypoglycaemia.

Cardiac arrhythmias and electrophysiologic responses during spontaneous hyperglycaemia in adults with type 1 diabetes mellitus.

AIM: We examined the effect of spontaneous hyperglycaemia in adults with type 1 diabetes mellitus (T1DM) and without history of cardiovascular disease on heart rate variability (HRV), cardiac repolarisation and incidence of cardiac arrhythmias. METHODS: Thirty-seven individuals with T1DM (age 17-50 years, 19 males, mean duration of diabetes 19.3 SD(9.6) years) underwent 96 h of simultaneous ambulatory 12-lead Holter ECG and blinded continuous interstitial glucose (IG) monitoring (CGM). HRV, QT interval and cardiac repolarisation were assessed during hyperglycaemia (IG ≥ 15 mmol/l) and compared with matched euglycaemia (IG 5-10 mmol/l) on a different day, separately during the day and night. Rates of arrhythmias were assessed by calculating incidence rate differences. RESULTS: Simultaneous ECG and CGM data were recorded for 2395 hours. During daytime hyperglycaemia vs euglycaemia the mean QTc interval duration was 404 SD(21)ms vs 407 SD(20)ms, P = 0.263. T-peak to T-end interval duration corrected for heart rate (TpTendc) shortened: 74.8 SD(16.1)ms vs 79.0 SD(14.8)ms, P = 0.033 and T-wave symmetry increased: 1.62 SD(0.33) vs 1.50 SD(0.39), P = 0.02. During night-time hyperglycaemia vs euglycaemia, the mean QTc interval duration was 401 SD(26)ms vs 404 SD(27)ms, P = 0.13 and TpTend shortened: 62.4 SD(12.0)ms vs 67.1 SD(11.8)ms, P = 0.003. The number of cardiac arrhythmias was low and confined to bradycardia and isolated ectopic beats. A considerable inter-subject and diurnal variability was observed. CONCLUSIONS: Hyperglycaemia in individuals with T1DM without known cardiovascular disease was not associated with clinically important cardiac arrhythmias.

The heart is a target organ for androgen.

Autoradiographic and biochemical analyses of the hearts of female rhesus monkeys and baboons indicate that atrial and ventricular myocardial cells contain androgen receptors. Although the specific effects of nuclear uptake and retention of androgen on the function of heart muscle cells are not known, the presence of this receptor suggests that sex steroid hormones may affect myocardial function directly and may explain some of the peculiar differences in heart disease between men and women.

Reduced leucine-enkephalin--like immunoreactive substance in hamster basal ganglia after long-term ethanol exposure.

Golden Syrian hamsters were placed individually in cages with three drinking bottles--one empty, one containing water, and the third containing water and ethanol. Control hamsters received water only. After 1 year the experimental hamsters showed a significantly lower concentration of leucine-enkephalin-like immunoreactive substance in the basal ganglia than the control hamsters. This finding indicates that the action of ethanol involves endogenous peptidyl opiates.

Androgen receptors in the brain: what are we measuring?

Androgen has long been known to act on the brain to modify behavior and other brain functions. In the past, two methods have been used to characterize the putative receptors which mediate these actions. Autoradiography has been used to map and identify androgen binding neurons. Binding studies have been conducted to quantify and characterize the system(s). The resultant data are discordant and a new model is proposed to resolve the apparent differences. It is proposed that there are three categories of receptors for androgen in the brain. One receptor preferentially binds testosterone and a second one preferentially binds DHT. Both of these receptors are in equilibrium between nucleus and cytoplasm according to the free water content of the compartments. Both of these receptors can be activated and transformed by steroid and thus concentrate in the nucleus. It is proposed that a third receptor binds both steroids with the same relative affinity. However, this third receptor can only be activated but not transformed (i.e. it does not concentrate in the nucleus). The proposed system implies that testosterone acts on a few discrete populations of neurons in the brain while DHT has a very diffuse action on the central nervous system.

Effect of zinc administration on the growth of L1210 and BW5147 tumors in mice.

Aqueous zinc acetate injected ip prevented tumor growth in 50-70% of BDF male mice previously inoculated ip with L1210 leukemia cells. However, aqueous zinc acetate injected sc did not prevent tumor growth in AKR/J mice inoculated im with BW5147 lymphatic leukemia cells. In the latter mice, only a small but statistically significant increase in mean survival was noted.

Allelic association of the D2 dopamine receptor gene with receptor-binding characteristics in alcoholism.

The allelic association of the human D2 dopamine receptor gene with the binding characteristics of the D2 dopamine receptor was determined in 66 brains of alcoholic and non-alcoholic subjects. In a blinded experiment, DNA from the cerebral cortex was treated with the restriction endonuclease Taql and probed with a 1.5-kilobase (kb) digest of a clone (lambda hD2G1) of the human D2 dopamine receptor gene. The binding characteristics (Kd [binding affinity] and Bmax [number of binding sites]) of the D2 dopamine receptor were determined in the caudate nuclei of these brains using tritiated spiperone as the ligand. The adjusted Kd was significantly lower in alcoholic than in nonalcoholic subjects. In subjects with the A1 allele, in whom a high association with alcoholism was found, the Bmax was significantly reduced compared with the Bmax of subjects with the A2 allele. Moreover, a progressively reduced Bmax was found in subjects with A2/A2, A1/A2, and A1/A1 alleles, with subjects with A2/A2 having the highest mean values, and subjects with A1/A1, the lowest. The polymorphic pattern of the D2 dopamine receptor gene and its differential expression of receptors suggests the involvement of the dopaminergic system in conferring susceptibility to at least one subtype of severe alcoholism.

Direct and quantitative detection of HIV-1 RNA in human plasma with a branched DNA signal amplification assay.

AIM: To determine the relative effect of sample matrix on the quantitation of HIV RNA in plasma. METHOD: Two HIV-positive specimens were diluted into five and 10 different HIV-negative plasma samples, respectively. Branched DNA signal amplification technology and reverse-transcriptase polymerase chain reaction were used to measure the viral load. RESULTS: In one sample the viral load by polymerase chain reaction ranged from undetectable to 1.9 x 10(5) copies/ml, and the branched DNA results ranged from 2.6 x 10(4) to 4.2 x 10(4) HIV RNA equivalent/ml. In the other sample the corresponding figures were 6.3 x 10(4) to 5.5 x 10(5) copies/ml and 5.7 x 10(4) to 7.5 x 10(4) HIV RNA equivalents/ml. CONCLUSION: In contrast to reverse-transcriptase polymerase chain reaction the branched DNA signal amplification assay does not require a separate extraction step or enzymatic amplification of the target. Therefore this measurement is less affected by the sample matrix and the signal generated is directly proportional to the viral load.

The primate spinal cord is a target for gonadal steroids.

The nuclear uptake and retention of 3H-dihydrotestosterone (3H-DHT) or one of its metabolites was studied in the spinal cord of the rhesus monkey. Normally-cycling adult female rhesus monkeys which were castrated and adrenalectomized prior to the experiment were injected with 1 microgram of 3H-DHT (107 Ci/mmole)/kg body weight and killed 90 minutes later. The spinal cords were removed and segments processed for autoradiography. Nuclear uptake and retention were found in both the visceral and somatic motor systems and, in addition, in the nociceptive system. The data suggest a role for androgen in sexual reflexes and possibly pain perception at the level of the spinal cord in the primate and provide further support for a role of androgen in amyotropic lateral sclerosis.

The nuclear uptake and retention of a synthetic progestin in the cardiovascular system of the baboon.

It has long been known that there is a sexual dimorphism in the incidence of coronary heart disease. This observation, together with more recent reports of increased cardiovascular disease associated with the use of oral contraceptives, led to a search for steroid receptors in the cardiovascular system. In this study we examined the nuclear uptake and retention of a synthetic progestin in the cardiovascular system of the baboons. Long term oophorectomized baboons were primed with estradiol benzoate for 3 days before the experiment (50 micrograms/kg, im) and adrenalectomized 2 days before the experiment. On the day of the experiment, the animals were injected under anesthesia with 2.5 micrograms/kg BW [3H]ORG 2058 (16 alpha-ethyl-21-hydroxy-19-nor-[6,7-3H]pregn-4-ene-3,20-dione) or with [3H] ORG 2058 plus a 1000-fold excess of unlabeled progesterone (control). One hour after the injection, the animals were rapidly exsanguinated, and parts of the cardiovascular system were removed and processed for autoradiography. Localization of the synthetic progestin was found in nuclei of between 25-75% of all smooth muscle cells of the media of all arteries examined and to a lesser extent in the nuclei of the fibroblasts and others cells of the adventitia. Localization of the synthetic progestin in the heart was limited to approximately 1% of the myocardial cells and less than 5% of interstitial cell nuclei. The pattern of localization found differs from that for estrogen and androgen and suggests the possible presence of estrogen-independent progesterone receptors in smooth muscle cells of the media of the aorta and coronary arteries.

Unbound progesterone receptors are in equilibrium between the nucleus and cytoplasm in cells of the rat uterus.

The classical model for the mechanism of action of steroids holds that unbound receptors for steroids reside exclusively in the cytoplasmic compartment and that they undergo translocation to the nucleus when bound to steroids in a process which is temperature sensitive. We have in the past proposed that unbound receptors for estrogen are in both nucleus and cytoplasm in a state of equilibrium. In the present study we looked at the location of the progesterone receptor using autoradiography and biochemical procedures. Uteri were incubated with [3H]progesterone or [3H]R5020 (dimethyl-19-nor-pregna-4,7-diene-3,20 dione, 17 alpha, 21-[17 alpha-methyl-3H] for 5 min at 4 C. When the tissue was processed for autoradiography, the localization of steroid was nuclear. In contrast, when the tissue was processed using the usual biochemical procedures, all binding activity appeared in the cytoplasm. In addition, when concentrated preparations of homogenized uteri were made, free receptor could be demonstrated in the crude nuclear preparations. We hypothesize that unbound progesterone receptor, like unbound estrogen receptor in the rat uterus, is in both the nucleus and cytoplasm of cells. In addition, we propose that the intracellular distribution of unbound receptors for all steroids is dependent upon the equilibrium conditions present.

The primate thyroid gland contains receptors for androgens.

The gonadal steroids have long been known to modulate thyroid function. Most studies suggest that the gonadal steroids act indirectly through the hypothalamic-pituitary axis to modulate thyroid function. The following studies were conducted to determine whether there are receptors for androgens in the thyroid itself. Cytosols from male and female euthyroid patients were analyzed for the presence of androgen with the synthetic analog methyltrienolone [( 3H]R1881). No evidence of androgen receptors was found in any of the cytosols prepared from female patients. In all males studied, androgen receptors were found in concentrations ranging from 100-150 fmol/10 mg DNA for the cytosols and from 690-2800 fmol/10 mg DNA for the nuclear extracts. The receptors had a dissociation constant (Kd) of approximately 5-10 X 10(-10) M for the cytosol and approximately 10-15 X 10(-10) M for the nuclear extracts. In addition to the human studies, studies in baboons were conducted to determine the possible cell type which might contain receptors for androgens. Male and female baboons were injected with [3H] dihydrotestosterone and killed between 1 and 1 1/2 h later. The thyroids were removed and processed for autoradiography. In autoradiograms from animals injected with [3H]dihydrotestosterone, nuclear localization of radioactivity was found in virtually all of the follicular cells. Also, label was found overlying the colloid, with heaviest labeling near the cells. These data suggest that there may be direct actions of androgens on follicular cells.

Prolonged P300 latency in a neuropsychiatric population with the D2 dopamine receptor A1 allele.

The role of the dopaminergic system in P300 has been implicated and previous studies have suggested the presence of a heritable component in the genesis of P300 or P3, a late positive component of the event-related potential. In the present investigation, 155 Caucasian male and female diagnosed neuropsychiatrically-ill patients with and without comorbid drug and alcohol abuse/dependence were genotyped for the presence or absence of the A1 allele of the D2 dopamine receptor gene (DRD2). The relationship of the A1 and A2 alleles to P3 amplitude and latency was also determined. The results showed no significant difference in P3 amplitude between all groups studied with A1 and A2 allele carriers. However, we now report prolonged P3 latency in neuropsychiatrically-ill patients (with or without polysubstance abuse) with those carrying two copies of the A1 allele (homozygote) of the DRD2 gene (quadratic trend, p = 0.01). Moreover, the age-adjusted mean P3 latency in the D2A2/A2 allele group was 327.8 +/- 3.08 ms compared by ANOVA, to 360.04 +/- 4.86 ms in the D2A1/A1 group. Our work suggests an association of polymorphisms of the DRD2 gene and a biological marker previously indicated to have predictive value in vulnerability to substance abuse.

Dopamine D2 receptor gene variants: association and linkage studies in impulsive-addictive-compulsive behaviour.

Drug and alcohol seeking behaviour has become a great global problem affecting millions of inhabitants with a cost to society in the billions. Dopaminergic reward pathways have frequently been implicated in the etiology of addictive behaviour. While other neurotransmitters have also been implicated, to date the only molecular genetic defect which has been found to associate with alcoholism, drug dependency, obesity, smoking, pathological gambling, attention-deficit-hyperactivity disorder (ADHD), Tourette syndrome, as well as other related compulsive behaviours, are the variants of the dopamine D2 receptor gene (DRD2). In this review of the available data on the subject, we report a number of independent meta-analyses that confirm an association of DRD2 polymorphisms and impulsive-additive-compulsive behaviour (IACB), which we have termed "Reward Deficiency Syndrome". While we agree that Meta-analyses of all exant studies support an association of variants of DRD2 and IACB, correct negative findings with alcoholism may be due to differences in assessing controls and inclusion/exclusion criteria for selection of diseased probands.

Increased prevalence of the Taq I A1 allele of the dopamine receptor gene (DRD2) in obesity with comorbid substance use disorder: a preliminary report.

In order to investigate the prevalence of the Taq I A1 allele of the dopamine receptor gene (DRD2) in obesity with and without comorbid substance use disorder, a total of 40 patients, from an outpatient neuropsychiatric clinic in Princeton, New Jersey, were genotyped for presence or absence of the Taq I DRD2 A1 allele. The primary inclusion criterion for 40 obese subjects was a body mass index (BMI) equal to or over 25 (uncharacterized); 11 obese subjects had severe substance use disorder; 20 controls had a BMI below 25; and, 33 substance use disorder (less severe) patients had a BMI below 25. The data were statistically compared with three different sets of controls divided into three separate groups (Group I, n = 20; Group II, n = 286; Group III, n = 714). They differed according to screening criteria (drug, alcohol, nicotine abuse/dependence, BMI below 25 and other related behaviours including parental history of alcoholism or drug abuse and DSM IV, Axis I and Axis II diagnoses). Groups II and III were population controls derived from the literature. The prevalence of the Taq I A1D2 dopamine receptor (DRD2) alleles was determined in 40 Caucasian obese females and males. In this sample with a mean BMI of 32.35 +/- 1.02, the A1 allele of the DRD2 gene was present in 52.5% of these obese subjects. Furthermore, we found that in the 23 obese subjects possessing comorbid substance use disorder, the prevalence of the DRD2 A1 allele significantly increased compared to the 17 obese subjects without comorbid substance use disorder. The DRD2 A1 allele was present in 73.9% of the obese subjects with comorbid substance use disorder compared to 23.5% in obese subjects without comorbid substance use disorder. Moreover, when we assessed severity of substance usage (alcoholism, cocaine dependence, etc.) increasing severity of drug use increased the prevalence of the Taq I DRD2 A1 allele; where 66.67% (8/12) of less severe probands possessed the A1 allele compared to 82% (9/11) of the most severe cases. Linear trend analyses showed that increasing use of drugs was positively and significantly associated with A1 allelic classification (p < 0.00001). These preliminary data suggest that the presence of the DRD2 A1 allele confirms increased risk not only for obesity, but also for other related addictive behaviours (previously referred to as the Reward Deficiency Syndrome) and that a BMI over 25 by itself (without characterization of macroselection or comorbid substance use disorders) is not a sufficient criterion for association with the DRD2 A1 allele.

Androgen-concentrating cells in the periventricular brain of the female rhesus monkey.

Although androgens act on the primate central nervous system to modulate both endocrine functions and a number of limbic-related behaviors, little is known about the anatomical location of the neurons which sequester these steroids in primates. To determine the prime location of these androgen-concentrating neurons in the forebrain of the primate, we injected three castrated female rhesus monkeys in the femoral vein with 1 microgram of 5 alpha-dihydro (1,2,4,5,6,7-3H) testosterone (3H-DHT, 107 Ci/mmole) per kg of body weight. One of these animals also received an IV injection of 100 micrograms/kg body weight of unlabeled dihydrotestosterone (DHT) to serve as a control. One hour after the injection of 3H-DHT we rapidly exsanguinated each animal. The forebrain was sliced and blocks containing the amygdala, diencephalon, frontal pole, and hippocampus were frozen and stored in liquid nitrogen until processing. The tissue was then processed for autoradiography. A specific topographic pattern of nuclear concentration of DHT or one of its metabolites was obtained in neurons of the basal hypothalamus, preoptic region, amygdala, and hippocampus. This pattern was similar to that found in rodent species.

Expression of three viral oncogenes (v-sis, v-myc, v-fos) in primary human brain tumors of neuroectodermal origin.

We determined which viral oncogenes (v-sis, v-myc, and v-fos) were expressed in five primary human brain tumors of neuroectodermal origin (two glioblastomas multiforme, one medulloblastoma, one cystic cerebellar astrocytoma, and one ganglioglioma) and which of these oncogenes is correlated with malignancy. Using the dot hybridization technique, we determined the relative amounts of mRNA coded by these genes using the same nitrocellulose filter. The v-myc probe showed a 4- to 12-fold greater hybridization to the mRNA from two glioblastomas and the medulloblastoma (malignant group) than the mRNA from the cystic cerebellar astrocytoma or the ganglioglioma (benign group). In contrast, RNA hybridizing to v-sis and v-fos were accumulated to a greater extent in the benign tumors. These data suggest that the amount of myc expression may be correlated with the degree of malignancy of brain tumors of neuroectodermal origin.

Autoradiographic and biochemical studies of hormone receptor localization.

Autoradiography has been used in the past to locate presumptive receptor systems for a number of steroids. It has provided information in rather complex tissues that could not have been obtained by biochemical procedures. In these studies we made use of autoradiography to redirect our biochemical efforts to study androgen receptors in the primate heart. Castrated-adrenalectomized female rhesus monkeys and baboons were injected with 1 microgram and 5 alpha-dihydro-[1,2,4,5,6,7-3H]-testosterone per kilogram of body weight. The animals were killed 1 hour later and the hearts were removed and processed for autoradiography. A nuclear localization of androgen was found in most of the myocardial cells, but in few if any of the interstitial cells. After reviewing the autoradiographic data, we began a new series of biochemical studies using a new buffer system and unlabeled androgen to stabilize the receptor. This in combination with postlabeled gradients allowed us to demonstrate for the first time 8S binding on sucrose density gradients. We feel that autoradiography can be a useful adjunct to biochemical studies even in "less complex" tissue such as the cardiac muscle.

Can a single androgen receptor fill the bill?

If the above two hypotheses are correct, they would require at least one more specific nuclear receptor for T, and at least one membrane receptor to account for the very rapid effects induced by androgens on certain target tissues. If this is the case, clearly a single androgen receptor will not fill the bill.

Medically induced gingival hyperplasia.

Gingival hyperplasia or gingival overgrowth is a common occurrence in patients taking phenytoin, cyclosporine, or calcium channel blockers. Speech, mastication, tooth eruption, and aesthetics may be altered. Controlling the inflammatory component through an appropriate oral hygiene program may benefit the patient by limiting the severity of the gingival overgrowth. In patients in whom gingival overgrowth is present or may be anticipated, recognition of this condition and referral to a general dentist or periodontist are appropriate steps to management. The physician's awareness of the potential for development of overgrowth and the dental practitioner's role in attempting to prevent or minimize this problem are important aspects. In this article, we discuss the medications associated with gingival hyperplasia and describe appropriate recommendations.