Plasma dopamine concentrations following dopamine infusion to isoflurane-anesthetized New Zealand White rabbits.

OBJECTIVE: To determine the pharmacokinetics of dopamine following a short infusion in isoflurane-anesthetized rabbits. STUDY DESIGN: Prospective, descriptive pharmacokinetic study. ANIMALS: A group of six adult female New Zealand White rabbits weighing 4.4 ± 0.2 kg. METHODS: Rabbits were anesthetized with isoflurane in oxygen and maintained at 1.2 × minimum alveolar concentration of isoflurane (2.3% atmosphere). Dopamine (30 µg kg-1 minute-1) was infused for 10 minutes. Arterial blood was sampled prior, during and following the infusion at various intervals for 1 hour. RESULTS: A one-compartment model with baseline concentration best fitted the time-plasma dopamine concentration data. Estimated typical population value (interindividual variability) for volume of distribution and clearance were 10.3 (232%) L kg-1 and 9.9 (508%) L minute-1 kg-1, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: There was a large degree of interindividual variation in the disposition of dopamine. The large volume of distribution and high metabolic clearance rate reported for dopamine in this study likely explains the lack of clinical efficacy of dopamine in rabbits at doses up to 30 µg kg-1 minute-1.

Analysis of endogenous epinephrine and norepinephrine enantiomers in rat plasma and application to a stereoselective pharmacokinetics.

Epinephrine and norepinephrine are a class of chiral endogenous catecholamines, which are known as major neurotransmitters. This work described a new LC-MS/MS method coupled with pre-column derivatization, enabling the simultaneous enantiomeric separation of epinephrine and norepinephrine in rat plasma. After protein precipitation procedure, the samples were derivatized with (S)-N-(4-nitrophenoxycarbonyl) phenylalanine methoxyethyl ester, [(S)-NIFE]. The derivatives resolved with good baseline separation on an ACQUITY UPLC BEH C18 column (100 mm × 2.1 mm, 1.7 µm) with mobile phase composed of methanol with 0.2% formic acid in water at a flow rate of 0.2 mL/min. Analysis was performed by multiple reaction monitoring in positive ionization mode. The linear ranges were 1.0-500 ng/mL for epinephrine enantiomers and 1.5-750 ng/mL for norepinephrine enantiomers. The lower limits of quantification for epinephrine and norepinephrine enantiomers were 1.0 and 1.5 ng/mL, respectively. The intra-day and inter-day precision were all less than 10.7% and accuracy ranged from 96.0 to 101.5%. Recoveries for all the analytes were more than 80.3%. The proposed method was successfully applied to simultaneously determine endogenous epinephrine and norepinephrine enantiomers in rat plasma. l-epinephrine and l-norepinephrine were sensitively and accurately quantified while both the d-enantiomers were not detected. Additionally, epinephrine enantiomers were analyzed for stereoselective pharmacokinetics in rats after intravenous administration of racemic epinephrine for the first time. The pharmacokinetic results indicated that the disposition of epinephrine enantiomers was stereoselective and chiral inversion did not occur in rats.

Pharmacokinetic effects of endotracheal, intraosseous, and intravenous epinephrine in a swine model of traumatic cardiac arrest.

INTRODUCTION: Limited prospective data exist regarding epinephrine's controversial role in managing traumatic cardiac arrest (TCA). This study compared the maximum concentration (Cmax), time to maximum concentration (Tmax), plasma concentration over time, return of spontaneous circulation (ROSC), time to ROSC, and odds of ROSC of epinephrine administered by the endotracheal (ETT), intraosseous (IO), and intravenous (IV) routes in a swine TCA model. METHODS: Forty-nine Yorkshire-cross swine were assigned to seven groups: ETT, tibial IO (TIO), sternal IO (SIO), humeral IO (HIO), IV, CPR with defibrillation (CPRD), and CPR only. Swine were exsanguinated 31% of their blood volume and cardiac arrest induced. Chest compressions began 2 min post-arrest. At 4 min post-arrest, 1 mg epinephrine was administered, and blood specimens collected over 4 min. Resuscitation continued until ROSC or 30 min elapsed. RESULTS: The Cmax of IV epinephrine was significantly higher than the TIO group (P = 0.049). No other differences in Cmax, Tmax, ROSC, and time to ROSC existed between the epinephrine groups (P > 0.05). Epinephrine levels were detectable in two of seven ETT swine. No significant difference in ROSC existed between the epinephrine groups and CPRD group (P > 0.05). Significant differences in ROSC existed between all groups and the CPR only group (P < 0.05). No significant differences in odds of ROSC were noted. CONCLUSIONS: The pharmacokinetics of IV, HIO, and SIO epinephrine were comparable. Endotracheal epinephrine absorption was highly variable and unreliable compared to IV and IO epinephrine. Epinephrine appeared to have a lesser role than volume replacement in resuscitating TCA.

Determination of d-amphetamine and diphenhydramine in beagle dog plasma by a 96-well formatted liquid-liquid extraction and capillary zone electrophoresis with field-amplified sample stacking.

This paper describes a method for quantification of d-amphetamine and diphenhydramine in beagle dog plasma by organic solvent field-amplified sample stacking (FASS)-capillary zone electrophoresis (CZE), using amlodipine as the internal standard. The separation was carried out at 25 °C in a 40.2 cm × 75 µm fused-silica capillary with an applied voltage of 20 kV using 25 mM phosphate-18.75 mM borate (pH 3.5). The detection wavelength was 200 nm. Clean-up and preconcentration of plasma biosamples were developed by 96-well formatted liquid- liquid extraction (LLE). In this study, the peak areas of d-amphetamine, diphenhydramine and amlodipine in the plasma sample increased by the factor of 48, 67 and 43 compared to the CZE without sample stacking. The method was suitably validated with respect to stability, specificity, linearity, lower limit of quantitation, accuracy, precision and extraction recovery. The calibration graph was linear from 2 to 500 ng/ml for d-amphetamine and 2-5000 ng/ml for diphenhydramine. All the validation data were within the required limits. Compared with the LC/MS/MS method that we previously established, there was no significant difference between the two methods in validation characteristics, except the LLOQs. The developed method was successfully applied to the evaluation of pharmacokinetic study of the Quick-Acting Anti-Motion Capsules (QAAMC) in beagle dogs.

An Infant with a Prolonged Sympathomimetic Toxidrome after Lisdexamfetamine Dimesylate Ingestion.

INTRODUCTION: Stimulant medications are approved to treat attention deficit hyperactivity disorder (ADHD) in children over the age of 6 years. Fatal ingestion of stimulants by children has been reported, although most ingestions do not result in severe toxicity. Lisdexamfetamine dimesylate, a once daily long-acting stimulant, is a prodrug requiring conversion to its active form, dextroamphetamine, in the bloodstream. Based on its unique pharmacokinetics, peak levels of d-amphetamine are delayed. We describe a case of accidental ingestion of lisdexamfetamine dimesylate in an infant. CASE REPORT: A previously healthy 10-month-old infant was admitted to the hospital with a 5-h history of tachycardia, hypertension, dyskinesia, and altered mental status of unknown etiology. Confirmatory urine testing, from a specimen collected approximately 16 h after the onset of symptoms, revealed an urine amphetamine concentration of 22,312 ng/mL (positive cutoff 200 ng/mL). The serum amphetamine concentration, from a specimen collected approximately 37 h after the onset of symptoms, was 68 ng/mL (positive cutoff 20 ng/mL). Urine and serum were both negative for methamphetamine, methylenedioxyamphetamine (MDA), methylenedioxymethamphetamine (MDMA, Ecstasy), and methylenedioxyethamphetamine (MDEA). During the hospitalization, it was discovered that the infant had access to lisdexamfetamine dimesylate prior to the onset of symptoms. CONCLUSION: Amphetamine ingestions in young children are uncommon but do occur. Clinicians should be aware of signs and symptoms of amphetamine toxicity and consider ingestion when a pediatric patient presents with symptoms of a sympathetic toxidrome even when ingestion is denied.

Characteristics of analytically confirmed 3-MMC-related intoxications from the Swedish STRIDA project.

BACKGROUND: 3-Methylmethcathinone (3-MMC) is a synthetic cathinone stimulant structurally related to the new psychoactive substance (NPS) mephedrone (4-methylmethcathinone, 4-MMC). We describe a case series of analytically confirmed intoxications involving 3-MMC presented to emergency departments in Sweden and included in the STRIDA project. STUDY DESIGN: Observational case series of consecutive patients with self-reported or suspected use of NPS presenting to hospitals in Sweden between August 2012 and March 2014. METHODS: NPS analysis was performed by a liquid chromatography-mass spectrometry (MS)/MS method that is updated with new substances as they appear. Data on clinical features were collected during Poisons Information Centre consultations and retrieved from medical records. RESULTS: 3-MMC was detected in 50 (6.4%) of the 786 cases included in the STRIDA project during the 20-month study period, with the peak occurring in August 2013. The age range of patients testing positive for 3-MMC was 17-49 years (median 24) and 76% of them were men. The 3-MMC concentration in serum ranged between 0.002 and 1.49 µg/mL (median, 0.091) and between 0.007 and 290 µg/mL (median, 3.05) in urine. Co-exposure to other NPS and/or traditional drugs was very common, and 3-MMC mono-intoxication was found in only 4 (8%) cases. The most frequent clinical features were tachycardia (48% of cases) and agitation (42%). Other features included a reduced level of consciousness (32%), dilated pupils (24%), hallucinations (20%), diaphoresis (12%), seizures (8%), and hyperthermia (6%). Most patients (60%) needed hospital care for only 1 day but in 8% for 3 days or longer. CONCLUSION: The majority of patients with analytically confirmed 3-MMC exposure had sympathomimetic features similar to those associated with mephedrone intoxication. However, the high incidence of co-exposure to other drugs makes the clinical interpretation difficult. Nevertheless, 3-MMC was associated with a high admittance rate to intensive care (30%), and detected in two cases with a fatal outcome, suggesting that 3-MMC is a harmful drug.

An evidence-based review of epinephrine administered via the intraosseous route in animal models of cardiac arrest.

OBJECTIVES: Intraosseous (IO) access, enabling the rapid administration of epinephrine during cardiac arrest (CA), is crucial in promoting optimal postresuscitation outcomes in patients with poor vascular access. There is a question whether IO-administered epinephrine is equivalent to intravenously administered epinephrine during CA. METHODS: The question guiding this evidence-based review was as follows: in adults suffering CA given epinephrine via the IO route, what is the resulting serum concentration of the drug compared to when administered intravenously? A search was conducted and the evidence appraised and leveled. RESULTS: Four animal studies met the inclusion criteria. The sources showed no definitive evidence supporting equivalence between intravenous and IO epinephrine administered during CA. Intravenously administered epinephrine provides increased and faster appearing serum concentrations than IO-administered epinephrine. Evidence indicated epinephrine given via the sternal IO route more closely approaches equivalence with intravenously administered epinephrine than when administered by the tibial IO route. CONCLUSIONS: The clinician should consider using proximal IO infusion sites such as the sternum or humerus when administering advanced cardiac life support drugs to rapidly achieve maximal therapeutic concentrations. Further studies are needed to determine the differences seen when epinephrine is administered by these routes during CA.

Comparison of tibial intraosseous, sternal intraosseous, and intravenous routes of administration on pharmacokinetics of epinephrine during cardiac arrest: a pilot study.

The purpose of this study was to determine and compare the maximum concentration (C(max)) and time to maximum concentration (T(max)) of epinephrine administered via tibial intraosseous (IO), sternal IO, and intravenous (i.v.) routes in a porcine model of cardiac arrest during cardiopulmonary resuscitation. Five pigs each were randomly assigned to 3 groups: tibial IO, sternal IO, and i.v. Cardiac arrest was induced with i.v. potassium chloride. After 2 minutes, cardiopulmonary resuscitation was initiated. Epinephrine was administered to each animal, and serial blood samples were collected over the next 3 minutes. Enzyme-linked immunosorbent assay was used to determine the epinephrine concentration. Multivariate analysis of variance helped determine if there were statistically significant differences between groups. There were significant differences in Cmax between the sternal IO and i.v. (P = .009) and tibial IO and i.v. (P = .03) groups but no significant difference between tibial and sternal IO groups (P = .75). Significant differences existed in Tmax between the tibial IO and i.v. (P = .04) and between tibial IO and sternal IO (P = .02) groups but no difference between the sternal IO and i.v. groups (P = .56). Intravenous administration of 1 mg of epinephrine resulted in a serum concentration 5.87 and 2.86 times greater than for the tibial and sternal routes, respectively.

Reverse-phase liquid chromatography with electrospray ionization/mass spectrometry for the quantification of pseudoephedrine in human plasma and application to a bioequivalence study.

A sensitive and selective reverse-phase liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) method was developed and validated to quantify pseudoephedrine (CAS 90-82-4) in human plasma. Phenacetin was used as the internal standard (I.S.). Sample preparation was performed with a deproteinization step using acetonitrile. Pseudoephedrine and I.S. were successfully separated using gradient elution with 0.5% trifluoroacetic acid (TFA) in water and 0.5% TFA in methanol at a flow-rate of 0.2 mL/min. Detection was performed on a single quadrupole mass spectrometer by a selected ion monitoring (SIM) mode via electrospray ionization (ESI) source. The ESI source was set at positive ionization mode. The ion signals of m/z 166.3 and 180.2 were measured for the protonated molecular ions of pseudoephedrine and I.S., respectively. The lower limit of quantification (LLOQ) of pseudoephedrine in human plasma was 10 ng/mL and good linearity was observed in the range of concentrations 10-500 ng/mL (R2 = 1). The intra-day accuracy of the drug containing plasma samples was more than 97.60% with a precision of 3.99-11.82%. The inter-day accuracy was 99.36% or more, with a precision of 7.65-18.42%. By using this analytical method, the bioequivalence study of the pseudoephedrine preparation was performed and evaluated by statistical analysis of the log transformed mean ratios of pharmacokinetic parameters. All the results fulfilled the standard criteria of bioequivalence, being within the 80-125% range which is required by the Korea FDA, US FDA, and EMEA to conclude bioequivalence. Consequently, the developed reverse-phase LC-ESI-MS method was successfully applied to bioequivalence studies of pseudoephedrine in healthy male volunteers.

Bruxism affects stress responses in stressed rats.

It has been proposed that suppression of stress-related emotional responses leads to the simultaneous activation of both sympathetic and parasympathetic divisions of the autonomic nervous system (ANS) and that the expression of these emotional states has a protective effect against ulcerogenesis. In the present study, we investigated whether stress-induced bruxism activity (SBA) has a physiological effect of on the stress-induced changes of the stomach, thymus, and spleen as well as blood leukocytes, cortisol, and adrenaline. This study demonstrated that SBA attenuated the stress-induced ulcer genesis as well as degenerative changes of thymus and spleen. SBA also attenuated increases of adrenaline, cortisol, and neutrophils in the blood. In conclusion, expression of aggression through SBA during stress exposure attenuates both stress-induced ANS response, including gastric ulcer formation.

Greater systemic lipolysis in women compared with men during moderate-dose infusion of epinephrine and/or norepinephrine.

Women have lower circulating catecholamine levels during metabolic perturbations, such as exercise or hypoglycemia, but similar rates of systemic lipolysis. This suggests women may be more sensitive to the lipolytic action of catecholamines, while maintaining similar glucoregulatory effects. The aim of the present study, therefore, was to determine whether women have higher rates of systemic lipolysis compared with men in response to matched peripheral infusion of catecholamines, but similar rates of glucose turnover. Healthy, nonobese women (n = 11) and men (n = 10) were recruited and studied on 3 separate days with the following infusions: epinephrine (Epi), norepinephrine (NE), or the two combined. Tracer infusions of glycerol and glucose were used to determine systemic lipolysis and glucose turnover, respectively. Following basal measurements of substrate kinetics, the catecholamine infusion commenced, and measures of substrate kinetics continued for 60 min. Catecholamine concentrations were similarly elevated in women and men during each infusion: Epi, 182-197 pg/ml and NE, 417-507 pg/ml. There was a significant sex difference in glycerol rate of appearance and rate of disappearance with the catecholamine infusions (P < 0.0001), mainly due to a significantly greater glycerol turnover during the first 30 min of each infusion: glycerol rate of appearance during Epi was only 268 +/- 18 vs. 206 +/- 21 micromol/min in women and men, respectively; during NE, only 173 +/- 13 vs. 153 +/- 17 micromol/min, and during Epi+NE, 303 +/- 24 vs. 257 +/- 21 micromol/min. No sex differences were observed in glucose kinetics under any condition. In conclusion, these data suggest that women are more sensitive to the lipolytic action of catecholamines, but have no difference in their glucoregulatory response. Thus the lower catcholamine levels observed in women vs. men during exercise and other metabolic perturbations may allow women to maintain a similar or greater level of lipid mobilization while minimizing changes in glucose turnover.

Sudden unexpected death in a female fitness athlete, with a possible connection to the use of anabolic androgenic steroids (AAS) and ephedrine.

The use of anabolic androgenic steroids (AAS) has been associated with different adverse effects, some of which potentially lethal. Most users of AAS are male, but the prevalence of such use appears to be increasing in females. Here we present a sudden unexpected death in a female fitness athlete with a possible connection to use of doping agents.

First case report of recreational use of 2,5-dimethoxy-4-chloroamphetamine confirmed by toxicological screening.

Routine toxicological screening is generally not undertaken in patients with recreational drug toxicity. We report here the benefits of toxicological screening in confirming drugs that have been ingested and potentially detecting drugs that have not previously been reported in the medical literature. In this case, the patient was reported to have ingested a combination of 2,5-dimethoxy-4-iodoamphetamine and methylenedioxymetamphetamine and developed sympathomimetic toxicity, but on extended toxicological screening he was shown to have actually ingested 2,5-dimethoxy-4-chloroamphetamine and methylenedioxymetamphetamine. As 2,5-dimethoxy-4-chloroamphetamine is a substituted amphetamine, it is covered under the generic Misuse of Drugs act (1971) in the UK; although in the majority of the EU it remains uncontrolled, as there is no similar generic drug legislation. We believe that discrepancies in the legal status of recreational drugs in the EU limit the effectiveness of drug enforcement policies and that EU drug legislation should be harmonized to ensure consistency.

Protracted 'pro-addictive' phenotype produced in mice by pre-adolescent phenylpropanolamine.

For decades, the sympathomimetic phenylpropanolamine (PPA; +/- -norepinephrine) was an active ingredient found in popular children's over-the-counter (OTC) cold, cough, and allergy medications. To examine the possibility that pre-adolescent PPA exposure may induce neuroadaptations that influence behavioral and neurochemical responding to cocaine, C57BL/6J mice were pretreated with PPA (0-40 mg/kg) during postnatal days 21-31. The behavioral and neurochemical responses to acute and repeated cocaine (4 x 15 mg/kg) were then assessed in adulthood when the mice were 10 weeks of age. Whereas pre-adolescent PPA exposure did not influence the acute locomotor response to 15 mg/kg cocaine, PPA pre-exposure dose-dependently enhanced the expression of cocaine-induced place conditioning, reduced the expression of locomotor sensitization, but did not influence cocaine-induced stereotypy. Pre-adolescent PPA exposure completely prevented the capacity of cocaine to elevate extracellular levels of catecholamines in the nucleus accumbens, but facilitated the development of cocaine-induced glutamate sensitization. Neither acute nor repeated cocaine altered extracellular GABA levels in the accumbens of control mice; however, 15 mg/kg cocaine lowered GABA levels by approximately 40% in PPA pretreated mice and this effect showed tolerance with repeated cocaine administration. These data provide the first evidence that early exposure to an OTC compound produces protracted effects upon cocaine-induced changes in nucleus accumbens neurotransmission that may contribute to a 'pro-addictive' phenotype in adulthood.

A novel method for oral stimulant administration in the neonate rat and similar species.

Sixty male and female Long-Evans hooded rats were administered 1, 2, or 5mg/kg methylphenidate (MPH) suspended in apple juice on postnatal day (P)15 or P40 using a novel, non-invasive oral administration technique. Plasma was collected 15 min after ingestion and analyzed by high performance liquid chromatography-mass spectrometry (HPLC-MS) to confirm appropriate concentrations. HPLC-MS plasma analysis showed levels comparable to previous gavage studies using MPH. We have used this method successfully in subsequent behavioral studies as well. Since therapeutic MPH in humans is typically administered orally, oral dosing methods that have been verified in the rodent model are of value. We recommend employment of this alternative oral dosing technique as it is minimally invasive, can be used anytime during postnatal development, and does not depend upon voluntary consumption.

Effects of elevated plasma adrenaline levels on substrate metabolism, effort perception and muscle activation during low-to-moderate intensity exercise.

The aim of this study was to differentiate the role of raised plasma adrenaline (Adr) concentrations from sympathoadrenal activation associated with moderate-intensity exercise, on muscle activation, cardiopulmonary responses, fuel metabolism, and ratings of perceived exertion (RPE) during low-intensity exercise. Two groups of subjects (MOD, n=6; LOW, n=7) cycled on two occasions for 90 min. MOD cycled at 68% VO(2max) with saline infusion, and at 34% VO(2max) with Adr infusion. LOW cycled twice at 34% VO(2max), with either Adr or saline infusion. Infusions (0.015 g Adr/kg/min) started at 15 min and increased plasma [Adr] somewhat higher than during exercise at 68% VO(2max) (approximately 1.9 vs. 1.4 nM, at 75 min). Mean plasma glucose and lactate concentrations during LOW were significantly higher with Adr than saline infusion (5.1+/-0.6 vs. 4.4+/-0.3 mmol/l, P<0.01 and 2.1+/-0.8 vs. 1.3+/-0.5 mmol/l, P<0.01, respectively). Elevated [Adr], without increased exercise intensity, did not alter glycogenolysis. There were also no effects of Adr infusion at 34% VO(2max) on heart rate, oxygen consumption, [FFA], respiratory exchange ratio, intramuscular triglyceride utilization, muscle activation or RPE. In conclusion, elevated [Adr] similar to those found during moderate-intensity exercise increased plasma glucose and lactate availability, but did not alter intramuscular fuel utilization, effort perception or muscle activation.

Feedback effects of circulating norepinephrine on sympathetic outflow in healthy subjects.

The amplitude of low-frequency (LF) oscillations of heart rate (HR) usually reflects the magnitude of sympathetic activity, but during some conditions, e.g., physical exercise, high sympathetic activity results in a paradoxical decrease of LF oscillations of HR. We tested the hypothesis that this phenomenon may result from a feedback inhibition of sympathetic outflow caused by circulating norepinephrine (NE). A physiological dose of NE (100 ng.kg(-1).min(-1)) was infused into eight healthy subjects, and infusion was continued after alpha-adrenergic blockade [with phentolamine (Phe)]. Muscle sympathetic nervous activity (MSNA) from the peroneal nerve, LF (0.04-0.15 Hz) and high frequency (HF; 0.15-0.40 Hz) spectral components of HR variability, and systolic blood pressure variability were analyzed at baseline, during NE infusion, and during NE infusion after Phe administration. The NE infusion increased the mean blood pressure and decreased the average HR (P < 0.01 for both). MSNA (10 +/- 2 vs. 2 +/- 1 bursts/min, P < 0.01), LF oscillations of HR (43 +/- 13 vs. 35 +/- 13 normalized units, P < 0.05), and systolic blood pressure (3.1 +/- 2.3 vs. 2.0 +/- 1.1 mmHg2, P < 0.05) decreased significantly during the NE infusion. During the NE infusion after PHE, average HR and mean blood pressure returned to baseline levels. However, MSNA (4 +/- 2 bursts/min), LF power of HR (33 +/- 9 normalized units), and systolic blood pressure variability (1.7 +/- 1.1 mmHg2) remained significantly (P < 0.05 for all) below baseline values. Baroreflex gain did not change significantly during the interventions. Elevated levels of circulating NE cause a feedback inhibition on sympathetic outflow in healthy subjects. These inhibitory effects do not seem to be mediated by pressor effects on the baroreflex loop but perhaps by a presynaptic autoregulatory feedback mechanism or some other mechanism that is not prevented by a nonselective alpha-adrenergic blockade.

Differing physiological effects of epinephrine in type 1 diabetes and nondiabetic humans.

Acute increases of the key counterregulatory hormone epinephrine can be modified by a number of physiological and pathological conditions in type 1 diabetic patients (T1DM). However, it is undecided whether the physiological effects of epinephrine are also reduced in T1DM. Therefore, the aim of this study was to determine whether target organ (liver, muscle, adipose tissue, pancreas, cardiovascular) responses to epinephrine differ between healthy subjects and T1DM patients. Thirty-four age- and weight-matched T1DM (n = 17) and healthy subjects (n = 17) underwent two randomized, single-blind, 2-h hyperinsulinemic euglycemic clamp studies with (Epi) and without epinephrine infusion. Muscle biopsy was performed at the end of each study. Epinephrine levels during Epi were similar in all groups (4,039 +/- 384 pmol/l). Glucose (5.3 +/- 0.06 mmol/l) and insulin levels (462 +/- 18 pmol/l) were also similar in all groups during the glucose clamps. Glucagon responses to Epi were absent in T1DM and significantly reduced compared with healthy subjects. Endogenous glucose production during the final 30 min was significantly greater during Epi in healthy subjects compared with T1DM (8.4 +/- 1.3 vs. 4.4 +/- 0.6 micromol.kg(-1).min(-1), P = 0.041). Glucose uptake showed almost a twofold greater decrease with Epi in healthy subjects vs. T1DM (Delta31 +/- 2 vs. Delta17 +/- 2 nmol.kg(-1).min(-1), respectively, P = 0.026). Glycerol, beta-hydroxybutyrate, and nonesterified fatty acid (NEFA) all increased significantly more in T1DM compared with healthy subjects. Increases in systolic blood pressure were greater in healthy subjects, but reductions of diastolic blood pressure were greater in T1DM patients with Epi. Reduction of glycogen synthase was significantly greater during epinephrine infusion in T1DM vs. healthy subjects. In summary, despite equivalent epinephrine, insulin, and glucose levels, changes in glucose flux, glucagon, and cardiovascular responses were greater in healthy subjects compared with T1DM. However, T1DM patients had greater lipolytic responses (glycerol and NEFA) during Epi. Thus we conclude that there is a spectrum of significant in vivo physiological differences of epinephrine action at the liver, muscle, adipose tissue, pancreas, and cardiovascular system between T1DM and healthy subjects.

5-HT1A and beta-adrenergic receptors regulate proliferation of rat blood lymphocytes.

Lymphocytes possess serotonin 5-HT(1A) and beta-adrenergic receptors, which have been related to cell proliferation. In the present report, lymphocytes of rat blood were isolated by Ficoll-Hypaque gradients and differential adhesion to plastic. They were cultured in RPMI medium for 72 h in the presence of the mitogens lipopolysaccharide concanavalin A and anti-CD3 antibody. The latter two stimulated the proliferation of lymphocytes, but not the first. Serotonin (0.1-100 microM) was added alone or in the presence of suboptimal concentrations of concanavalin A (2 microg/ml) or anti-CD3 antibody (0.4 microg/ml). The 5-HT(1A) receptor agonists, 8-hydroxy-2-(di-n-propylamino)tetralin and buspirone (0.1-100 microM) were also tested in the cultures. Serotonin, 8-hydroxy-2-(di-n-propylamino)tetralin and buspirone neither had any effect by themselves, nor modified the proliferation induced by the mitogens. Noradrenaline (25-1,000 microM) and the beta-adrenergic receptor agonist, isoproterenol (5-100 microM), produced a reduction of the activation induced by concanavalin A or anti-CD3 antibody in a dose-dependent manner. Increasing serotonin concentrations reduced the inhibitory effect of noradrenaline (300 microM). Variable concentrations of 8-hydroxy-2-(di-n-propylamino)tetralin or buspirone also reduced the inhibition produced by isoproterenol (100 microM). The antagonist of 5-HT(1A) receptors, WAY-100,478 (0.1-100 microM), inhibited concanavalin A- or anti-CD3 antibody-induced proliferation. Serotonin (0.1-100 microM) impaired the inhibitory effect of the 5-HT(1A) antagonist (10 microM). The inhibitor of tryptophan hydroxylase, p-chlorophenylalanine (50-1,000 microM), decreased the stimulatory effect of concanavalin A, serotonin (0.5-100 microM) and 8-hydroxy-2-(di-n-propylamino)tetralin (1-100 microM) reverted the effect of p-chlorophenylalanine (1,000 microM). The serotonin reuptake blockers zimelidine, imipramine and clomipramine decreased concanavalin A-induced proliferation. The concentrations of serotonin and noradrenaline increased in lymphocytes cultured in the presence of concanavalin A, probably as a mechanism for modifying the final effect on proliferation. The present results indicate that 5-HT(1A) receptors play a stimulatory role on rat blood lymphocytes, and they interact in a parallel and opposite manner with beta-adrenergic receptors. Furthermore, endogenous serotonin is relevant in displaying its stimulatory effect.