Antagonist-induced Increase Research Articles

196-OR: Pharmacological Modulation of Prostaglandin E2 Receptor Activity Enhances ß-Cell Mass in a Mouse Model of Type 2 Diabetes and Human Islets

Type 2 diabetes (T2D) is characterized by chronic hyperglycemia and low-grade inflammation. Failure to expand β-cell mass in response to insulin resistance can lead to T2D. The hyperglycemia and chronic inflammation observed in T2D is associated with enhanced levels of prostaglandin E2 (PGE2) synthesis in pancreatic islets. Downstream effects of PGE2 signaling are carried out by four E-Prostanoid (EP) G-protein coupled receptors, EP1-4. Recent investigations by our lab demonstrated that EP3 and EP4 play opposing roles in pancreatic islets whereby EP3 activation inhibits β-cell proliferation and survival while EP4 activation stimulates β-cell mass expansion and reduces cytokine-induced cell death ex vivo. Preliminary data suggests that db/db mice (a model of T2D) treated with the EP3 antagonist, DG-041, exhibit enhanced levels of β-cell proliferation when compared to their vehicle-treated counterparts (3.04 ± 0.6% Ki67+/Ins+ β-cells and 1.67 ± 0.2%, respectively). Although differences in β-cell death between the vehicle- (0.09 ± 0.1% TUNEL+ β-cells) and DG-041-treated cohorts (0.23 ± 0.1%) did not reach statistical significance, treatment with the EP3 antagonist ultimately culminated in improved β-cell mass in db/db mice (1.57 ± 0.2 mg) compared to vehicle-treated controls (0.94 ± 0.2 mg). Additionally, preliminary results suggest that the EP3 antagonist-induced increase in β-cell proliferation in nondiabetic human cadaver donors occurs, in part, through PLCγ-1 whereby pretreatment of human islets with U-73122 (PLCγ-1 inhibitor) prior to DG-041 addition resulted in decreased β-cell proliferation (1.67 ± 0.2 fold of vehicle-treated control) when compared to the EP3 antagonist alone (2.30 ± 0.3 fold). Taken together, these results provide valuable foundational insight into the potential role of EP3 in the modulation of β-cell mass expansion. Disclosure S.R. Andrei: None. J.C. Dunn: None. A.A. Christensen: None. V.F. Ricciardi: None. W.A. Pace: None. R.M. Breyer: None. M. Gannon: None.

Dopamine D1 receptor activation maintains motor coordination in injured rats but does not accelerate the recovery of the motor coordination deficit

The sensorimotor cortex and the striatum are interconnected by the corticostriatal pathway, suggesting that cortical injury alters the striatal function that is associated with skilled movements and motor learning, which are functions that may be modulated by dopamine (DA). In this study, we explored motor coordination and balance in order to investigate whether the activation of D1 receptors (D1Rs) modulates functional recovery after cortical injury. The results of the beam-walking test showed motor deficit in the injured group at 24, 48 and 96h post-injury, and the recovery time was observed at 192h after cortical injury. In the sham and injured rats, systemic administration of the D1R antagonist SCH-23390 (1mg/kg) alone at 24, 48, 96 and 192h significantly (P<0.01) increased the motor deficit, while administration of the D1R agonist SKF-38393 alone (2, 3 and 4mg/kg) at 24, 48, 96 and 192h post-injury did not produce a significant difference; however, the co-administration of SKF-38393 and SCH-23390 prevented the antagonist-induced increase in the motor deficit. The cortical+striatal injury showed significantly increased the motor deficit at 24, 48, 96 and 192h post-injury (P<0.01) but did not show recovery at 192h. In conclusion, the administration of the D1R agonist did not accelerate the motor recovery, but the activation of D1Rs maintained motor coordination, confirming that an intact striatum may be necessary for achieving recovery.

Genetic and pharmacological inhibition of retinoic acid receptor γ function promotes endochondral bone formation.

The nuclear retinoic acid receptors (RARs) play key roles in skeletal development and endochondral ossification. Previously, we showed that RARγ regulates chondrogenesis and that pharmacological activation of RARγ blocked heterotopic ossification (HO), pathology in which endochondral bone forms in soft tissues. Thus, we reasoned that pharmacological inhibition of RARγ should enhance endochondral ossification, leading to a potential therapeutic strategy for bone deficiencies. We created surgical bone defects in wild type and RARγ-null mice and monitored bone healing. Fibrous, cartilaginous, and osseous tissues formed in both groups by day 7, but more cartilaginous tissue formed in mutants within and around the defects compared to controls. Next, we implanted a mixture of Matrigel and rhBMP2 subdermally to induce ectopic endochondral ossification. Administration of RARγ antagonists significantly stimulated ectopic bone formation in wild type but not in RARγ-null mice. The antagonist-induced increases in bone formation were preceded by increases in cartilage formation and were accompanied by higher levels of phosphorylated Smad1/5/8 (pSmad1/5/8) compared to vehicle-treated control. Higher pSmad1/5/8 levels were also observed in cartilaginous tissues forming in healing bone defects in RARγ-null mice, and increases in pSmad1/5/8 levels and Id1-luc activity were observed in RARγ antagonist-treated chondrogenic cells in culture. Our data show that genetic or pharmacological interference with RARγ stimulates endochondral bone formation and does so at least in part by stimulating canonical BMP signaling. This pharmacologic strategy could represent a new tool to enhance endochondral bone formation in the setting of various orthopedic surgical interventions and other skeletal deficiencies. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1096-1105, 2017.

Serotoninergic and dopaminergic modulation of cortico-striatal circuit in executive and attention deficits induced by NMDA receptor hypofunction in the 5-choice serial reaction time task

Executive functions are an emerging propriety of neuronal processing in circuits encompassing frontal cortex and other cortical and subcortical brain regions such as basal ganglia and thalamus. Glutamate serves as the major neurotrasmitter in these circuits where glutamate receptors of NMDA type play key role. Serotonin and dopamine afferents are in position to modulate intrinsic glutamate neurotransmission along these circuits and in turn to optimize circuit performance for specific aspects of executive control over behavior. In this review, we focus on the 5-choice serial reaction time task which is able to provide various measures of attention and executive control over performance in rodents and the ability of prefrontocortical and striatal serotonin 5-HT1A, 5-HT2A, and 5-HT2C as well as dopamine D1- and D2-like receptors to modulate different aspects of executive and attention disturbances induced by NMDA receptor hypofunction in the prefrontal cortex. These behavioral studies are integrated with findings from microdialysis studies. These studies illustrate the control of attention selectivity by serotonin 5-HT1A, 5-HT2A, 5-HT2C, and dopamine D1- but not D2-like receptors and a distinct contribution of these cortical and striatal serotonin and dopamine receptors to the control of different aspects of executive control over performance such as impulsivity and compulsivity. An association between NMDA antagonist-induced increase in glutamate release in the prefrontal cortex and attention is suggested. Collectively, this review highlights the functional interaction of serotonin and dopamine with NMDA dependent glutamate neurotransmission in the cortico-striatal circuitry for specific cognitive demands and may shed some light on how dysregulation of neuronal processing in these circuits may be implicated in specific neuropsychiatric disorders.

Effect of acute and chronic vitamin D administration on systemic renin angiotensin system in essential hypertensives and controls

To investigate the systemic renin-angiotensin system (RAS) in essential hypertensives (EH) and controls (C) after short- and long-term vitamin D receptor activation. Ten consecutive EH (under controlled low-salt diet) and 10 C underwent calcitriol administration (0.25 μg bid) for 1 week (Group A). Eighteen consecutive EH under angiotensin II receptor antagonist therapy received a single oral dose of 300,000 IU of cholecalciferol and were followed up for 8 weeks (Group B). In basal conditions and at the end of the study (1 week in Group A and 8 weeks in Group B), plasma renin activity (PRA), plasma active renin, aldosterone, and angiotensin II were evaluated, as well as blood pressure, plasma 25-hydroxyvitamin D [25(OH)D], 1,25-dihydroxyvitamin D [1,25(OH)2D], and PTH. In Group A, plasma 25(OH)D levels in EH and C were below the normal range, although lower levels were found in the former. No association between basal plasma 25(OH)D or 1,25(OH)2D levels and blood pressure values or RAS components was observed either in the whole group or in the two subgroups. Calcitriol administration did not affect any RAS parameter either in EH or in C. In Group B, cholecalciferol significantly increased 25(OH)D and 1,25(OH)2D levels without interfering with the angiotensin II receptor antagonist-induced increase in RAS components. No correlation was found between plasma 25(OH)D or 1,25(OH)2D levels and blood pressure values or RAS parameters before and after cholecalciferol administration. The present data suggest that, in our experimental conditions, vitamin D receptor activation is unable to influence systemic RAS activity.

Evidence for involvement of nitric oxide and GABAB receptors in MK-801- stimulated release of glutamate in rat prefrontal cortex

Systemic administration of NMDA receptor antagonists elevates extracellular glutamate within prefrontal cortex. The cognitive and behavioral effects of NMDA receptor blockade have direct relevance to symptoms of schizophrenia, and recent studies demonstrate an important role for nitric oxide and GABAB receptors in mediating the effects of NMDA receptor blockade on these behaviors. We sought to extend those observations by directly measuring the effects of nitric oxide and GABAB receptor mechanisms on MK-801-induced glutamate release in the prefrontal cortex. Systemic MK-801 injection (0.3 mg/kg) to male Sprague–Dawley rats significantly increased extracellular glutamate levels in prefrontal cortex, as determined by microdialysis. This effect was blocked by pre-treatment with the nitric oxide synthase inhibitor l-NAME (60 mg/kg). Reverse dialysis of the nitric oxide donor SNAP (0.5–5 mM) directly into prefrontal cortex mimicked the effect of systemic MK-801, dose-dependently elevating cortical extracellular glutamate. The effect of MK-801 was also blocked by systemic treatment with the GABAB receptor agonist baclofen (5 mg/kg). In combination, these data suggest increased nitric oxide formation is necessary for NMDA antagonist-induced elevations of extracellular glutamate in the prefrontal cortex. Additionally, the data suggest GABAB receptor activation can modulate the NMDA antagonist-induced increase in cortical glutamate release.

Targeting Hyponatremia and Hemodynamics in Acute Decompensated Heart Failure: Is There a Role for Vasopressin Antagonists?

Current treatment of acute decompensated heart failure (ADHF) has not reduced the significant morbidity or mortality associated with this disease, and has promoted drug development aimed at neurohormonal targets. Hypervolemic hyponatremia, which is linked to the nonosmotic release of arginine vasopressin, is associated with a poor prognosis in patients with heart failure (HF). Vasopressin acts on V(2) and V(1a) receptors to cause water retention and vasoconstriction, respectively. Clinical trials have demonstrated that vasopressin receptor antagonists (VRAs) are effective in treating hypervolemic hyponatremia in ADHF without a negative impact on renal function. The small hemodynamic benefit seen with VRA use appeared to result from V(2)-receptor antagonist-induced increase in urine output rather than from a vasodilatory drug effect. VRA use in ADHF trials was associated with minimal symptomatic improvement and no impact on morbidity or mortality. At present, clinical trial evidence does not support the routine use of VRAs in ADHF. Given the favorable renal profile of VRAs, studies on the possible benefit of VRAs in ADHF patients with renal insufficiency and diuretic resistance appear warranted.

Retrograde Endocannabinoid Signaling in a Postsynaptic Neuron/Synaptic Bouton Preparation from Basolateral Amygdala

Retrograde synaptic signaling by endogenous cannabinoids (endocannabinoids) is a recently discovered form of neuromodulation in the brain. In the basolateral amygdala (BLA), endocannabinoid signaling has been implicated in learning and memory, specifically in extinction of aversive memories. To examine retrograde endocannabinoid signaling in this brain region, BLA neurons were freshly isolated using an enzyme-free procedure. These isolated neurons retain attached functional excitatory and inhibitory synaptic boutons. Spontaneous GABAergic IPSCs (sIPSCs) were isolated from these freshly isolated neurons and a 4 s step of depolarization from -60 to 0 mV produced suppression of sIPSC frequency and amplitude. A similar depolarization-induced suppression of inhibition (DSI) was observed in neurons in BLA slices. DSI in the single-cell preparation was abolished by the CB1 receptor antagonist N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide, and DSI duration was shortened in the presence of 2-methyl-6-(phenylethynyl) pyridine, an mGluR5 (metabotropic glutamate receptor 5) antagonist. The initial decrease in sIPSCs induced by the DSI procedure was greatly attenuated in recordings with 20 mm BAPTA containing postsynaptic internal solution, but a delayed-onset decrease was observed under this recording condition. A CB1 agonist decreased sIPSC frequency and amplitude, whereas CB1 antagonists increased these responses. The antagonist-induced increase was abolished in 20 mm BAPTA-filled cells. These data provide solid evidence for retrograde endocannabinoid signaling in the BLA and also indicate that this retrograde signaling requires only a postsynaptic neuron and attached synaptic boutons.

Modulation of noradrenaline release in the median preoptic area by GABAergic inputs from the organum vasculosum of the lamina terminalis in the rat

Previous observations have shown that γ-aminobutyric acid (GABA) receptor mechanisms modulate the release of noradrenaline (NA) in the median peptic nucleus (MnPO). The present study was carried out to investigate whether neural inputs from the organum vasculosum of the lamina terminalis (OVLT) to the MnPO are involved in the GABAergic modulation of NA release in the MnPO area using in vivo microdialysis techniques. In urethane-anesthetized rats, electrical stimulation (5 and 10 μA, 10 Hz) of the OVLT region, but not its surrounding region, significantly enhanced dialysate NA concentration in the MnPO area. The enhancement in the NA level caused by the OVLT region stimulation was significantly increased by perfusion with either bicuculline (10 μM), a GABA A receptor antagonist, or phaclofen (10 μM), a GABA B receptor antagonist, through a microdialysis probe. The amount of the antagonist-induced increase was much greater in the phaclofen-treated group than in the bicuculline-treated group. These results show that the OVLT region may exert both excitatory and inhibitory influences on the release of NA in the MnPO area, and imply that the inhibitory influence may be mediated through GABA B receptors rather than GABA A receptors.

Characterization of the distribution of the cocaine priming threshold and the effect of SCH23390

The cocaine-induced reinstatement (priming) of cocaine self-administration occurs when the cumulative concentration of cocaine reaches a threshold level that we have previously termed the cocaine priming threshold. The present studies used a modified procedure to measure the cocaine priming threshold over 4–8-month periods in individual rats. The values for the priming threshold varied between days but there was no evidence of a systematic change in the priming threshold over time, indicating that neither tolerance nor sensitization occurred. The frequency distribution of the priming threshold was significantly different from a normal distribution but was not significantly different from a log-normal distribution. Therefore, the geometric mean with its associated variance estimates, but not the arithmetic mean, appropriately describe the distribution of the cocaine priming threshold. The estimate of the geometric mean value of the priming threshold for this group of Sprague–Dawley rats was 284 (CI 95: 234–344) μg/kg of cocaine. The log-normal distribution of equieffective doses of cocaine is typical of agonist-induced pharmacological responses. In the presence of the D 1 dopamine receptor-selective antagonist SCH23390, the geometric means of the cocaine priming threshold were significantly increased in a dose-dependent manner, implying a role for D 1 dopamine receptors in the priming response. This technique provides a quantitative method for the measurement of antagonist-induced increases in the cocaine priming threshold.

Gamma Vinyl-GABA Differentially Modulates NMDA Antagonist-Induced Increases in Mesocortical Versus Mesolimbic DA Transmission

To explore the role of endogenous GABA in NMDA antagonist induced dopamine (DA) release, we used in vivo microdialysis to study the effects of pretreatment with γ-vinyl GABA (GVG) on phencyclidine (PCP)-induced DA release in terminal regions of midbrain DA neurons. GVG, an irreversible inhibitor of the GABA catabolizing enzyme GABA-AT, significantly reduced the DA response to PCP (7.0 mg/kg) in freely moving animals. Preferential increases in PCP-induced DA release in the PFC (four-fold those of NAcc) were dose-dependently inhibited by acute pretreatment with GVG at doses of 150 (51% inhibition), 300 (68% inhibition), and 500 (82% inhibition) mg/kg, whereas NAcc PCP-induced DA activity was unresponsive to 150 mg/kg and only partially inhibited by 300 and 500 mg/kg. Subchronic treatment with GVG did not enhance the inhibitory capacity of the GABAergic system. While GVG evidently modulates PCP-induced increases in mesocorticolimbic DA transmission, the character of this modulation is regionally specific, with cortical NMDA-antagonist induced increases appearing more sensitive to inhibition by endogenous GABA than subcortical areas.

Adrenocorticosteroid receptor blockade and excitotoxic challenge regulate adrenocorticosteroid receptor mRNA levels in hippocampus.

The mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) are glucocorticoid-activated transcription factors essential for maintenance of cellular homeostasis. Differential activation of these adrenocorticosteroid receptors (ACR) is thought to influence neuronal viability, particularly under challenging cellular conditions. The present study is designed to determine the effects of receptor blockade and excitotoxic insult on MR and GR mRNA expression and neuronal viability in hippocampus. Male Sprague--Dawley rats were pretreated for 48 hr with vehicle, MR antagonist spironolactone (SPIRO) (50 mg/kg, twice daily, s.c.), or GR antagonist RU486 (25 mg/kg, twice daily, s.c.) and subsequently injected with saline or the glutamate analog kainic acid (KA) (12 mg/kg i.p.). Twenty-four hr post-insult, MR and GR mRNA levels were assessed by in situ hybridization analysis, and hippocampal neurons were counted to assess KA-induced cell loss. MR blockade with SPIRO increased basal MR mRNA levels in hippocampal subregions CA1, CA3, and dentate gyrus (DG) and increased basal GR mRNA levels in CA3. GR blockade with RU486 increased basal GR mRNA levels in CA3. The excitotoxin KA decreased MR mRNA levels in CA1 and CA3, decreased GR mRNA levels in DG, and negated all antagonist-induced increases of ACR mRNAs. Cell counts quantifying KA damage indicated increased CA3 vulnerability to KA insult after treatment with MR antagonist spironolactone but demonstrated no significant cell loss in any other group or region. These results demonstrate dynamic regulation of hippocampal MR and GR mRNAs after ACR antagonist treatment and kainate toxicity, underscoring the potential importance of MR and GR availability to neuronal viability after insult.

Role of the astrocytic ETB receptor in the regulation of extracellular endothelin‐1 during hypoxia

Astrocytes are known to possess an effective endothelin (ET) eliminatory system which involves astrocytic ET(A) and ET(B) receptors and may become particularly relevant under pathophysiological conditions. The present study has therefore been designed to explore the effect of standardized hypoxia on extracellular concentrations of endothelin-1 (ET-1) and on endothelin-converting enzyme (ECE) activity in primary rat astrocytes genetically (sl/sl) or experimentally (dexamethasone) deficient in ET(B) receptors. The results revealed (1) a hypoxia-mediated decrease of extracellular ET-1 in wildtype astrocytes (+/+) that was not observed in ET(B)-deficient (sl/sl) cultures; (2) an ET receptor antagonist-induced increase in ET-1 in the media of both genotypes with further elevation upon hypoxia in +/+ cultures only; (3) augmentation of the dexamethasone-induced increase in extracellular ET-1 by hypoxia in +/+, but not in sl/sl cultures; (4) synergistic reduction of ET(B) gene transcription by hypoxia and dexamethasone; and (5) significant increases in endothelin-converting enzyme activity in the presence of hypoxia. To conclude, hypoxia stimulates astrocytic release of mature ET-1. This stimulation is (over)compensated for by increased ET-1 binding to functional ET(B) receptors. ET(B) deficiency, whether genetic or experimentally induced, impairs elimination of extracellular ET-1.

Quantitative immunolocalization of mu opioid receptors: regulation by naltrexone

The present study utilized a newly developed quantitative immunohistochemical assay to measure changes in mu opioid receptor abundance following chronic administration of the opioid receptor antagonist naltrexone. These data were compared with those obtained from mu receptor radioligand binding on adjacent tissue sections, in order to determine whether the characteristic antagonist-induced increase in radioligand binding is due to an increase in the total number of mu receptors and/or to an increase in the proportion of receptors that are in an active binding conformation in the absence of a change in the total number of receptors. Adult male Sprague–Dawley rats were administered naltrexone, 7–8 mg/kg per day, or saline continuously for seven days by osmotic minipumps, after which time their brains were processed for immunohistochemistry and receptor autoradiography on adjacent fresh frozen tissue sections. Semiquantitative immunohistochemistry was performed using a radiolabelled secondary antibody for autoradiographic determination and a set of radioactive standards. Results demonstrate an overall concordance between the distribution of mu opioid receptors as measured by the two different methods with a few exceptions. Following naltrexone administration, mu receptor immunoreactivity was significantly higher in the amygdala, thalamus, hippocampus, and interpeduncular nucleus as compared with the saline-treated control animals. [ 3H]D-Ala 2,N-Me-Phe 4,Gly-ol 5-enkephalin binding to mu opioid receptors was significantly higher in the globus pallidus, amygdala, thalamus, hypothalamus, hippocampus, substantia nigra, ventral tegmental area, central gray, and interpeduncular nucleus of the naltrexone-treated rats. These findings indicate that in some brain regions chronic naltrexone exposure increases the total number of mu opioid receptors, while in other regions there is an increase in the percent of active receptors without an observable change in the total number of receptors. Quantitative receptor immunodetection together with ligand autoradiography provides a new approach for investigating the regulation of mu opioid receptors on tissue sections.

The role of GABA in modulating the Xenopus electroretinogram.

We have recorded the electroretinogram (ERG) from the superfused eyecup of the Xenopus retina in order to assess the effects of the inhibitory neurotransmitter gamma-aminobutyric acid (GABA), and its agonists and antagonists, on individual ERG components. We found that GABA (0.5-10 mM) reduced the amplitudes of both the b- and d-waves of the Xenopus ERG. The GABA uptake blocker nipecotic acid (1 mM) had similar effects on b- and d-waves. GABA at 5 mM and 10 mM also caused an increase in the a-wave. The GABA antagonist picrotoxin (0.1-2 mM) and the GABA/a antagonist bicuculline (0.2 mM) both increased the amplitude of the b- and d-waves of the ERG. The GABA/b agonist baclofen (0.3 mM) reduced the amplitude of the ERG b-wave, enhanced the amplitude of the a-wave, and slightly reduced the amplitude and increased the peak time of the d-wave. The GABA/b antagonists phaclofen and saclofen had no reliable effects on the Xenopus ERG. Glutamate analogs known to affect specific types of retinal neurons were applied to modify the retinal circuitry and then the effects of GABA and its antagonists were examined under these modified conditions. 2-amino-4-phosphonobutyric acid (APB) increased the d-wave, and blocked the b-wave and the effect of GABA on the ERG, but not the antagonist-induced increase in the d-wave. KYN blocked the antagonist-induced increase in the b-wave, while GABA increases the amplitude of the b-wave if the d-wave has been removed by prior superfusion with kynurenic acid (KYN). N-methyl-DL-aspartate (NMDLA), which acts only in the proximal retina, reduced the amplitude of the ERG and blocked the effect of GABA and the antagonist-induced increase in ERG b- and d-waves amplitude. These results suggest that GABAergic mechanisms related to both A and B receptor types can influence the amplitude and light sensitivity of all the components of the Xenopus ERG. Since GABA is found in greatest abundance in the proximal retina, and B type of receptors are present almost exclusively there, the data suggests that most of the effects of GABA agonists and antagonists observed are dependent on proximal retinal mechanisms, and that there are separate mechanisms in the proximal retina related to the b- and the d-waves.

Effects of calcium antagonists on binding of local anesthetics to plasma proteins and erythrocytes in mice.

This study was designed to document possible changes in the binding of bupivacaine (BV), lidocaine (LC) and their main metabolites desbutylbupivacaine (PPX) and monoethylglycinexylydide (MEGX), respectively, to plasma proteins and erythrocytes in mice after acute treatment with the calcium antagonists diltiazem (DZ), nicardipine (NP) and verapamil (VP). A significant plasma protein binding of BV, LC and PPX was measured, whereas no binding could be detected for MEGX. The binding of BV was not modified by DZ, NP and VP, however, the total plasma level was increased in the presence of VP. For PPX a significant increase in total and free plasma levels and a decrease in protein binding were demonstrated after DZ and VP treatment. Concerning LC, a significant increase in total and free plasma levels was documented for DZ, NP and VP suggesting an inhibition of the metabolism of LC by the calcium antagonists. An increased penetration of LC into erythrocytes was also demonstrated which is consistent with the calcium antagonist-induced increase in LC free plasma levels. These effects may contribute in part to the previously observed increase in toxicity of BV by calcium antagonists, but are not likely to be the sole mechanism.

Dopamine antagonist effects on behavior maintained by cocaine and alfentanil in rhesus monkeys.

The effects of dopamine (DA) antagonists that act on either the D1 site (SCH 39166), the D2 site (eticlopride), or both sites non-selectively (cis-flupenthixol) were evaluated for their effects on behavior maintained by cocaine or alfentanil in rhesus monkeys. Each of these drugs suppressed rates of responding maintained by cocaine or alfentanil. Larger doses of each of the DA antagonists were necessary to suppress cocaine- as opposed to alfentanil-maintained responding, suggesting that cocaine but not alfentanil was able to antagonize the rate-suppressing effects of the antagonists. There was little evidence, under these conditions of acute administration, that the DA antagonists modified the reinforcing effects of either cocaine or alfentanil. This would have been observed by an antagonist-induced increase in the ED(50) of the reinforcing drugs and, although such an increase was seen occasionally with cocaine, it was never statistically significant. The effects of rate-suppressing doses of each of the antagonists on directly observable behavior indicated a rapid onset and relatively short duration of action of intravenously administered SCH 39166 and eticlopride. cis-Flupenthixol had a much slower onset of action. Each of the DA antagonists produced similar increases in measures of sedation and relaxation. These data suggest very similar behavioral effects of DA antagonists that act selectively on D1 or D2 receptors or act non-selectively on both DA receptors.

Regulation of substance P receptor system in rat striatum by chronic naltrexone treatment

Chronic blockade of opioid receptors by naltrexone increases opioid peptides in the striatum, and up-regulates brain opioid receptors resulting in functional supersensitivity. Striatal SP content was increased 3.5-fold after 8 days of naltrexone treatment relative to control animals. The present study was undertaken to determine whether SP receptors in the striatum and SP receptor-coupled second messenger system are modulated by increased striatal SP content induced by chronic opioid receptor blockade. The binding affinity and capacity of SP receptors, determined using [ 125I]Bolton-Hunter SP ([ 125I]BHSP) labeled at Lys 3, in striatal synaptosomal membranes were not significantly altered by chronic blockade of opioid receptors. Although the concentrations of [Sar 9,Met (O 2) 11]SP, a NK-1 receptor-specific agonist, and SP(1–7), an aminoterminal major metabolite of SP, required to inhibit half of [ 125I]BHSP binding (IC 50) in striatal synaptosomal membranes were significantly decreased, the IC 50s for SP and an NK-2 receptor-specific agonist, [Nle 10]NK A(4–10), remained unchanged by chronic naltrexone treatment. The data suggest that naltrexone which has no SP receptor antagonistic action, not only indirectly acts on SP-ergic neurons but also causes a change in the apparent affinity of NK-1 receptor (as reflected by changes in IC 50 values) in the striatum. Cellular inositol-1,4,5-trisphosphate [Ins(1,4,5)P 3], quantified by a highly sensitive and selective radioreceptor mass assay, was increased in the striatum by 28% relative to control levels. With [ 3H]Ins(1,4,5)P 3 as a ligand, Scatchard analyses of the concentration-dependent saturation curves showed that the density of striatal intracellular Ins(1,4,5)P 3 receptors was increased by 53%. The levels of SP and cellular Ins(1,4,5)P 3, and the density of Ins(1,4,5)P 3 receptors in the cerebellum, used as a positive control, were unchanged by chronic naltrexone treatment. The findings of opiate antagonist-induced increases in SP striatal content and Ins(1,45)P 3 receptor densities, appear to support the concept of a role of endogenous opioids in the regulation of SP receptor activity. The data also suggest that inter-regulatory mechanisms exist between phospholipase C/phosphoinositide-coupled receptors such as SP receptors, and adenylate cyclase-coupled inhibitory receptors, such as opioid receptors.

Vasectomy modulates the local effect of intratesticular opioids on the gonad of immature rats.

Previous studies indicate that 7 days after testicular administration of naloxone, serum testosterone (T) concentration and basal T secretion in vitro decrease significantly. In neonatal rats, short-term (2 h) intratesticular treatment with 0.3 micrograms (D-Met2-Pro5)-enkephalinamide suppresses steroidogenesis. In the present study, testicular treatment with naloxone or enkephalinamide was combined with hemivasectomy (which also includes transection of the inferior spermatic nerve). When local treatment with naloxone was combined with vasectomy in 15-day-old rats, the opioid antagonist-induced increase in testicular weight, the decrease in basal T secretion of the treated gonad and the drop in serum T level could not be observed 7 days postsurgery, despite the fact that in immature rats, hemivasectomy, by itself, also reduces steroid secretion. Similarly, in 5-day-old animals, the short-term (2 h) effect of testicular enkephalinamide on T secretion was prevented by vasectomy. These data suggest that local testicular actions exerted by endogenous opioid peptides might be modulated by the inferior spermatic nerve.