Role of food-derived opioid peptides in regulation of thermosensitivity of <Emphasis Type="Italic">Periplaneta americana</Emphasis> cockroach

The ability of exorphines (relative δ-selective opioid fragments of food-derived peptides) to change the defense response of American cockroaches Periplaneta americana placed in a hot chamber (T= 50°C) was studied. It was found that the ED₅₀ for a fragment of D-ribulose-1,5-bisphosphate carboxylase rubiscolin-5 (YPLDL) doubling the stay time for the insects in the hot chamber was 340 nM/g. The ED₅₀ for wheat gluten exorphine C (YPISL) was 573 nM/g. Cytochropin-4, a fragment of cytochrome B, did not have a significant effect. Comparison of the obtained data with the results of previous experiments with μ-selective exorphines—milk β-casein fragments—led to the conclusion that opioid receptors of various types are equally involved in regulation of cockroach protective behavior. The significance of phytogenous exorphines in formation of the opioid system of insects in the course of evolution is discussed.     

Effects of Peptide and Non-Peptide Opioids on Protective Reaction of the Cockroach Periplaneta americana in the “Hot Camera”

The ability of morphine, naloxone, and several opioid peptides of the group of beta-casomorphines to change the time of the stay of cockroaches Periplaneta americana in a hot camera (t = 47°C) was studied. It has been shown that the morphine dose ED₅₀ increasing twice the stay amounts to 200 µg/g, while hat of naloxone, to 40, of heptapeptide YPFPGPI, to 440, and of pentapeptide YPFPG, to 420 µg/g. Hexapeptide YPFPGP free of the N-terminal tyrosine had no statistically significant effect on the stay duration. The earlier changes of the stay duration (in 15–60 min after injection; the most pronounced for morphine and naloxone) corresponded to the ability of these drugs to act on the mu-type opioid receptors. The high peptide affinity to the delta-type receptors led to development of the later effects (in 90–150 min after injection; the most pronounced for heptapeptide YPFPGPI). A combined injection of naloxone and heptapeptide lead to the mutual inhibition of their effect: the peptide eliminated the early effect of naloxone on the stay duration, whereas naloxone, the late effects of beta-casomorphine. The obtained results indicate an important role of the endogenous opioid system in control of protective behavior of insects, as well as heterogeneity of the receptor components of the system.     

Structure-activity relationship of rubiscolins as delta opioid peptides

To study the structure-activity relationship of rubiscolins (YPLDLF and YPLDL), delta opioid peptides derived from the spinach Rubisco, we substituted the amino acid residues and evaluated their activities by mouse vas deferens (MVD) and guinea pig ileum (GPI) assays as well as receptor affinity. Replacement of Leu(3) with Ile and Met in rubiscolin-6 potentiated the delta opioid activity by about four times in MVD assay. Asp(4) cannot be replaced by Ala, Glu or His. The original Leu(5) was optimal, while substitution of Phe(6) with Val potentiated its delta opioid activity by more than 10 times. The most potent derivative we obtained was YPMDLV, which was nearly 20 times more potent than rubiscolin-6 in MVD assay. The derivatives thus obtained showed higher delta receptor affinity and more potent antinociceptive activity than rubiscolins.     

In Vivo Characterization of (−)(−)MCL-144 and (+)(−)MCL-193: Isomeric,Bivalent Ligands with Mu/Kappa Agonist Properties

Once opioid receptor dimers were postulated, a goal has been to synthesize and screen novel opioids, with the hope of furthering our knowledge of the structure-activity relationship of opioid ligands with the opioid receptors. The aim of the current study was to address whether two isomeric bivalent ligands would have pharmacological differences after central administration, in vivo. The two compounds, (−) bis(N-cyclobutylmethyl-morphinan-3-yl) sebacoylate dihydrochloride (MCL-144) and 1−((+)N-cyclobutylmethylmorphinan-3-yl)-10-((−) N-cyclobutylmethylmorphinan-3-yl)sebacolyate (MCL-193) are each linked by a 10-carbon chain ester. The active (−) enantiomer for both ligands is 3-hydroxy-N-cyclobutylmethyl morphinan ((−)MCL-101), a N-cyclobutylmethyl analogue of cyclorphan (J Med Chem 43:114–122, 2000). MCL-144 contains two active levo rotatory (−)(−) pharmacophores, while MCL-193 contains one active (−) and one inactive (+) pharmacophore of MCL-101. In vitro analysis demonstrated that all three compounds, (−)(−)MCL-144, (+)(−)MCL-193 and (−)MCL-101 were κ agonists and μ partial agonists. (−)(−)MCL-144 and (−)MCL-101 had much higher affinity for both the μ and κ opioid receptors compared to (+)(−)MCL-193. In vivo, (−)(−)MCL-144 and (+)(−)MCL-193 produced full dose–response curves, in the 55°C tail-flick test, with each compound having an ED₅₀ value of 3.0 nmol after intracerebroventricular (i.c.v.) administration. The analgesic properties of both compounds were antagonized by the μ-selective antagonist, β-funaltrexamine and the κ-selective antagonist nor-binaltorphimine. Concomitant, i.c.v., administration of either (−)(−)MCL-144 or (+)(−)MCL-193 with morphine, did not significantly antagonize morphine-induced antinociception at any dose tested. In antinociceptive tests, (−)(−)MCL-144 and (+)(−)MCL-193 had the same pharmacological properties, demonstrating that having two active pharmacophores separated by a 10-carbon spacer group did not increase the antinociceptive efficacy of the compound. Additionally, it was also of interest to compare (−)(−)MCL-145 and (−)(−)MCL-144, as the only difference between these bivalent ligands is the spacer region connecting the two pharmacophores, yet (−)(−)MCL-145 produced an ED₅₀ value 10-fold lower than (−)(−)MCL-144 (ED₅₀ values = 0.3 nmol and 3.0 nmol, respectively). Special issue article in honor of Dr. Ji-Sheng Han.     

Anion modulation of the slowly activating vacuolar channel

A series of n-alkanols and phenyl-substituted n-alkanols (Φ-alkanols) of increasing chain length and phenol were characterized for their ability to block action potentials (APs) in frog sciatic nerves. APs were recorded using the single sucrose-gap method. The degree of AP attenuation when the nerve was exposed to different concentrations of an alcohol was used to construct dose-response curves. The reciprocals of the half-blocking doses (ED₅₀s) were used to obtain a measure of the potency of the alcohols. For n-alkanols and Φ-alkanols, increasing the chain length by the addition of a methylene group increased the potency on average by 3.1 for both groups of alkanols. The addition of a phenyl group caused a potency increase that ranged between the values of 77 and 122. The ED₅₀ for both groups of alkanols could not be solely predicted by the log octanol-water partition coefficient (K _OW ). Using linear solvation energy relations (LSER), the log ED₅₀ could be described as a linear combination of the intrinsic (van der Waals) molar volume (V _I ), polarity (P), and hydrogen bond acceptor basicity (β) and donor acidity (α). Size alone could not predict the ED₅₀ for both n-alkanols and Φ-alkanols. The results are consistent with the hypothesis that alkanols bind to and interact with Na channels to cause AP block. Phenyl group addition to an alkanol markedly increases the molecule's potency. Received: 11 August 2000/Revised: 21 December 2000     

Sigma1 binding in a human neuroblastoma cell line

Behaviorally, sigma₁ agents modulate opioid analgesia. To examine possible mechanisms responsible for these interactions, we have identified a cell line containing both sigma₁ and opioid receptors. [³H](+)-pentazocine binding in BE(2)-C human neuroblastoma cells is high affinity (K_D 3.4±0.7 nM) and high density (B_max 2.98±0.14 pmol/mg protein). Competition studies reveal a selectivity profile similar to that of sigma₁ sites in guinea pig brain. (+)-Pentazocine has no effect upon either basal or forskolin-stimulated cyclase in the BE(2)-C cells, but cAMP accumulation is inhibited by the morphine, DPDPE and naloxone benzoylhydrazone. (+)-Pentazocine at concentrations as high as 10 μM does not affect this opioid effect, implying that sigma₁/opioid interactions are not mediated at the level of the cell. This suggests that their behavioral interactions result from interacting neural circuits. Although (+)-pentazocine is without effect in the cyclase system, it does block carbachol-stimulated phosphoinositol turnover (IC₅₀ 6.5±1.14 μM). The specificity of the effect is confirmed by the ability of haloperidol (1 μM) to shift the IC₅₀ value of (+)-pentazocine 2-fold to the right. Special issue dedicated to Dr. Eric J. Simon.     

Cyanide inhibition and pyruvate-induced recovery of cytochrome <Emphasis Type="Italic">c</Emphasis> oxidase

The mechanism of cyanide’s inhibitory effect on the mitochondrial cytochrome c oxidase (COX) as well as the conditions for its recovery have not yet been fully explained. We investigated three parameters of COX function, namely electron transport (oxygen consumption), proton transport (mitochondrial membrane potential Δψ _m) and the enzyme affinity to oxygen (p ₅₀ value) with regard to the inhibition by KCN and its reversal by pyruvate. 250 μM KCN completely inhibited both the electron and proton transport function of COX. The inhibition was reversible as demonstrated by washing of mitochondria. The addition of 60 mM pyruvate induced the maximal recovery of both parameters to 60–80% of the original values. When using low KCN concentrations of up to 5 μM, we observed a profound, 30-fold decrease of COX affinity for oxygen. Again, this decrease was completely reversed by washing mitochondria while pyruvate induced only a partial, yet significant recovery of oxygen affinity. Our results demonstrate that the inhibition of COX by cyanide is reversible and that the potential of pyruvate as a cyanide poisoning antidote is limited. Importantly, we also showed that the COX affinity for oxygen is the most sensitive indicator of cyanide toxic effects.     

Cytoprotective and antisecretory properties of a non-diarrheogenic and non-uterotonic prostacyclin analog: U-68, 215

U-68,215 [15-Cyclohexyl-9-deoxo-13, 14-dihydro-2′, 9a-methano-4,5,6,16,17,18,19,20-octanor-3-oxa-3,7-(′1,′3-interphenylene)-PGE₁] is a stable prostacyclin analog. When given orally to rats, it is cytoprotective for the stomach (ED₅₀: 0.8 μg/kg) and the intestine (ED₅₀: 22 μg/kg), is gastric antisecretory (ED₅₀: 35 μg/kg) and antiulcer (aspirin) (ED_{50: 5 μg/kg}). The oral antisecretory ED₅₀ in dogs in 50 μg/kg. It has a long duration of gastric cytoprotection: 8–10 hours compared to 3 hours for 16, 16-dimethyl PGE₂ Unlike most prostaglandins of the E type, it is not diarrheogenic (not enteropooling), it does not induced cellular proliferation of the gastroeintestinal mucosa, when given twice a day for eight days, it is not uterotonic (in monkeys), and it does not prevent embryo implantation in hamsters. It ihibits platelet aggregation (ED₅₀: 300 μg/kg), but does not promote bleeding from cut vessels nor from gastric ulcers. U-68,215 lowers blood pressure at a n oral dose correponding to 1–5 time the antisecretory ED₅₀ in rats and dogs, and to 150 times the cytoprotective ED₅₀ in rats. It may be of therapeutic value in the treatment of conditions where inhibition of gastric acid secretion is desirable, e.g., gastric and duodenal ulcer, and in conditions responding to cytoprotection, e.g., stress ulcers, hemorrhagic gastritis and gastric erosions associated with nonsteroidal antiinflammatory drugs.     

Effects of Gabapentin on Different Neuronal Populations in the Dorsal-Root Ganglia of Rats with Streptozotocin-Induced <Emphasis Type="Italic">Diabetes Mellitus</Emphasis>

We studied the effect of gabapentin (an agent similar, in its molecular structure, to gamma-aminobutyric acid, GABA) on depolarization-evoked calcium transients in small, mid-sized, and large (diameter of the soma up to 25, 25 to 35, and 35 μm or more, respectively) neurons of the dorsal-root ganglia (DRGs) of rats with experimental streptozotocin-induced diabetes mellitus. These transients were measured using a calcium-sensitive fluorescent dye, Fura 2/AM. The amplitude of calcium transients in rats with diabetes was somewhat higher than that in healthy animals (in large and mid DRG neurons by nearly 12% and in small cells by about 8%, on average). The development of diabetes led to a dramatic increase in the total duration of such transients. In large, mid, and small DRG neurons, the values of this parameter in animals with diabetes were, respectively, about 260, 430, and 250% as compared with the norm. The duration of transients at the level of 50% amplitude (Т _0.5) in diabetes changed to a significantly smaller extent. Applications of gabapentin (25 μM) led to a decrease in the amplitude of calcium transients, their full duration, and Т _0.5. The effects of gabapentin were the strongest in large DRG neurons where the amplitude of calcium transients dropped by nearly 36%, while the total duration demonstrated a more than threefold decrease. Upon the action of gabapentin, the parameter Т _0.5 changed moderately (in all groups of DRG neurons, the decrease varied from 8 to 12%). Gabapentin-induced decreases in the amplitude of calcium transients differed in various subgroups of DRG neurons. Among neurons with mid-sized somata, the decrease in this parameter in capsaicin-positive cells was 16.3%, while that in capsaicin-negative cells reached 36.7%. The obtained data are indicative of the ability of gabapentin to normalize, to a certain extent, the parameters of diabetes-modified calcium transients in DRG neurons. This ability is more clearly pronounced in large neurons (we hypothesize that a part of such cells in animals with diabetes are, probably, abnormally involved in transmission of nociceptive influences) and also in a part of mid-sized DRG neurons participating in the formation of acute pain sensation.     

TAPP analogs containing β3‐homo‐amino acids: synthesis and receptor binding

β‐Amino acids containing α,β‐hybrid peptides show great potential as peptidomimetics. In this paper, we describe the synthesis and affinity to μ‐opioid and δ‐opioid receptors of α,β‐hybrids, analogs of the tetrapeptide Tyr‐ d ‐Ala‐Phe‐Phe‐NH₂ (TAPP). Each amino acid was replaced with an l ‐ or d ‐β³‐h‐amino acid. All α,β‐hybrids of TAPP analogs were synthesized in solution and tested for affinity to μ‐opioid and δ‐opioid receptors. The analog Tyr‐β³h‐ d ‐Ala‐Phe‐PheNH₂ was found to be as active as the native tetrapeptide. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.     

Synthesis and receptor binding of opioid peptide analogues containing β3‐homo‐amino acids

β‐Amino acids containing hybrid peptides and β‐peptides show great potential as peptidomimetics. In this paper we describe the synthesis and affinity toward the µ‐ and δ‐opioid receptors of β‐peptides, analogues of Leu‐enkephalin, deltorphin I, dermorphin and α,β‐hybrides, analogues of deltorphin I. Substitution of α‐amino acid residues with β³‐homo‐amino acid residues, in general resulted in decrease of affinity to opioid receptors. However, the incorporation β³h‐D ‐Ala in position 2 or β³hPhe in position 3 of deltorphin I resulted in potent and selective ligand for δ‐opioid receptor. The NMR studies of β‐deltorphin I analogue suggest that conformational motions in the central part of the peptide backbone are partially restricted and some conformational preferences can be expected. Copyright © 2009 European Peptide Society and John Wiley & Sons, Ltd.     

Potentiation of intrathecal DAMGO antinociception,but not gastrointestinal transit inhibition,by 5-hydroxytryptamine and norepinephrine uptake blockade

Spinally administered μ opioid agonists produce potent antinociception and inhibition of gastrointesdtinal transit. Blockade of 5-hydroxytryptamine (5-HT) or norepinephrine (NE) uptake potentiates intrathecal (i.t.) DAMGO antinociception. To determine whether 5-HT and NE uptake blockade will also potentiate the gastrointestinal inhibition, mice were treated with zimelidine, desipramine or saline, followed by i.t. DAMGO and tested for tail-flick antinociception or inhibition of gastrointestinal transit. DAMGO produceed antinociception dose-dependently (ED₅₀ = 4.6 ng). Zimelidine (10 mg/kg, s.c., 1 hr before DAMGO) produced a 6.2-fold ng). Desipramine produced a 5.3-fold shift (ED₅₀ = 1.4 ng). DAMGO also produced a dose-dependent inhibition of gastrointestinal transit (ED₅₀ = 117 ng). However, zimelidine or desipramine treatment did not affect DAMGO inhibition of gastrointestinal transit (ED₅₀ = 80 ng.).     

Effects of cyclic adenosine 3′:5′-monophosphate-elevating agents and retinoic acid on differentiation in retinoid-deficient tracheal cultures

Summary The ability of cyclic AMP-elevating agents to induce normal differentiation has been investigated in retinoid-deficient hamster tracheal epithelium in organ culture. Dibutyryl cAMP (dbcAMP) and other cAMP-regulating agents alone caused disappearance of keratin and regeneration of normal mucociliary epithelium in retinoid-deficient cultures. Incubation of retinoid-deficient cultures with dbcAMP, isoproterenol, and cholera toxin (CT) (without addition of exogenous retinoid) reversed keratinization in a dose-dependent manner. The ED₅₀ of cultures treated with dbcAMP was 4×10⁻⁶ M; ED₅₀ of isoproterenol was 7×10⁻⁵ M; and CT, 0.6 μg/ml. Phosphodiesterase inhibitors and other cAMP analogs were inactive. Dibutyryl cAMP in combination with theophylline enhanced normal differentiation. Retinoid-deficient tracheas pretreated for 20 h with 10⁻⁹ M all-trans-retinoic acid (RA) responded to 10⁻⁶ M dbcAMP by potentiating normal differentiation; this concentration of dbcAMP alone was inactive. Isoproterenol showed a similar response but to a lesser degree. These cAMP-elevating agents applied in combination with theophylline did not increase activity. This investigation was supported by National Cancer Institute Contract NO1-CP-31012.     

Stimulation of the adenylyl cyclase activity in human endometrial membranes by VIP and related peptides

Vasoactive intestinal peptide (VIP) has been shown to stimulate adenylyl cyclase activity in human endometrial membranes. The effect was dependent on the time and temperature of incubation as well as on the concentration of endometrial membrane proteins in the medium. In the presence of 1 M GTP, half-maximal stimulation of adenylyl cyclase activity was observed at 25.0±7.0 nM VIP, whereas the maximal activity (at 1 M VIP)corresponded to an increase of about 140% with respect to basal values (7.5±0.6 pmol cyclic AMP/min/mg of protein). However, the maximal stimulation of adenylyl cyclase activity was obtained with helodermin (1 M) that increased the activity by 170% over the basal. The relative potency of VIP-related peptides upon the adenylyl cyclase activity was: helodermin (ED₅₀=1.8±1.4 nM)>VIP(ED₅₀=25.0±7.0 nM)>PHI (ED₅₀=725.0±127.2 nM). Secretin had a faint effect upon the adenylyl cyclase activity and glucagon was completely inefficient at this level. The presence of _s and _i subunits of G proteins in human endometrium was detected by immunoblot. Preliminary results showed the presence of two classes of¹²⁵I-VIP receptors in human endometrial membranes with the following stoichoimetric parameters: high affinity receptor (Kd=2.0 nM, binding capacity 0.1 pmol VIP/mg protein) and low affinity receptor (Kd=0.43 M, binding capacity 13.1 pmol VIP/mg protein). The present results together with the known presence of VIP in human uterus and the actions of this neuropeptide in the adjacent myometrial tissue support the idea that VIP and related peptides may have a role in human endometrium.     

The influence of chronic anaemia on catecholamine secretion in the rainbow trout (Oncorhynchus mykiss)

The effect of long-term (7 day) anaemia on catecholamine release was examined in rainbow trout (Oncorhynchus mykiss) in vivo during acute exposure to hypoxia and in situ using a perfused post-cardinal vein preparation. The first goal was to distinguish among reductions in blood O₂ partial pressure, O₂ concentration and haemoglobin percentage saturation as potential stimuli for, or correlates of, catecholamine secretion during hypoxia. The second goal was to elucidate the role of these factors in promoting enhanced chromaffin cell responsiveness in trout subjected to chronic hypoxia (Montpetit and Perry 1998). Anaemic fish (haematocrit lowered from 28.4±2.4% to 11.9±1.6%) displayed a marked reduction in haemoglobin-O₂ binding affinity [P ₅₀ (P _aO₂ at 50% Hb-O₂ saturation) was increased from 14.7 mm Hg to 24.3 mm Hg]. Upon exposure to hypoxia, the anaemic fish released catecholamines into their circulation at higher values of arterial O₂ partial pressure (∼52 mm Hg versus ∼18 mm Hg) and haemoglobin O₂ saturation (<70% versus <55%) than did control fish. In addition, anaemic fish achieved significantly greater circulating levels of total catecholamines (noradrenaline plus adrenaline) during acute hypoxia (294.8±67.3 versus 107.0±35.6 nmol l⁻¹). These results do not support the view that catecholamine release is triggered by a reduction in haemoglobin O₂ saturation or arterial PO₂, per se. Nor are they consistent with the idea that catecholamine release occurs at a threshold value of arterial PO₂ corresponding to a critical reduction in blood O₂ concentration. The effects of the non-selective cholinergic receptor carbachol on catecholamine secretion from chromaffin tissue were assessed using perfused posterior cardinal vein preparations derived from control or anaemic fish. For adrenaline secretion, there was no statistically significant change in the ED₅₀ (dose eliciting 50% response). For noradrenaline secretion however, preparations originating from anaemic fish displayed an enhanced responsiveness to carbachol as indicated by a significant 4.5-fold reduction in the carbachol ED₅₀ value from 2.53 × 10⁻⁶ mol kg⁻¹ to 5.67 × 10⁻⁷ mol kg⁻¹. These results demonstrate that anaemia-induced hypoxaemia, in the absence of any lowering of PO₂, is able to modulate the responsiveness of chromaffin cells to cholinergic stimulation. Accepted: 21 April 1999     

The influence of exogenous peptide on β2–microglobulin exchange in the HLA complex: analysis in real-time

 We used an optical biosensor to determine the relative binding affinity of peptides to purified HLA class I molecules. In this assay we monitor β₂–microglobulin (β₂m) exchange within the HLA-A2 molecule, whereby native β₂m in the complex is replaced by β₂m immobilized at the surface of the biosensor. Quantitative kinetic measurements permit us to obtain association rate (k_ass), dissociation rate (k_diss) and affinity constants (K_A) for the β₂m exchange reaction, alone, (control) and in the presence of exogenous peptide. We tested a panel of six peptides which had been designed and synthesized with an HLA-A2 binding motif, and had also been tested by the T2-cell binding assay, along with control peptides. The biosensor results demonstrate that exogenous peptide influences the dynamics of β₂m exchange in a sequence-specific manner. Five of six peptides increased the association rate, decreased the dissociation rate, and significantly increased the affinity (K_A=1.55–1.88×10⁹ M^–1) of HLA-A2 for immobilized β₂m compared with the control (K_A =1.14±0.04×10⁹ M^–1), demonstrating stabilization of the complex. One peptide was unable to stabilize the complex, as also shown in the T2 binding assay. However, analysis of peptide sequences demonstrated that the HLA-A2 secondary motif as well as primary motif residues are required for HLA-A2 stabilization. Further experiments demonstrated that β₂m exchange alone cannot stabilize the HLA class I complex at the cell surface until a peptide of sufficient binding affinity is bound. Hence kinetics equal to or below the control values in our biosensor assay probably represent an unstable complex in vivo. Unlike other methods described for the analysis of peptide stabilization, this approach is significantly faster, provides full kinetic analysis, and is simpler, since it requires no labeling of peptides. Furthermore, this may have important implications in the assessment of peptide vaccines. Received: 9 October 1997 / Revised: 20 January 1998     

Cariprazine (RGH-188), a potent D3/D2 dopamine receptor partial agonist, binds to dopamine D3 receptors in vivo and shows antipsychotic-like and procognitive effects in rodents

We investigated the in vivo effects of orally administered cariprazine (RGH-188; trans-N-{4-[2-[4-(2,3-dichlorophenyl)-piperazin-1-yl]-ethyl]-cyclohexyl}-N′,N′-dimethyl-urea), a D₃/D₂ dopamine receptor partial agonist with ∼10-fold preference for the D₃ receptor. Oral bioavailability of cariprazine at a dose of 1 mg/kg in rats was 52% with peak plasma concentrations of 91 ng/mL. Cariprazine 10 mg/kg had good blood-brain barrier penetration, with a brain/plasma AUC ratio of 7.6:1. In rats, cariprazine showed dose-dependent in vivo displacement of [³H](+)-PHNO, a dopamine D₃ receptor-preferring radiotracer, in the D₃ receptor-rich region of cerebellar lobules 9 and 10. Its potent inhibition of apomorphine-induced climbing in mice (ED₅₀ = 0.27 mg/kg) was sustained for 8 h. Cariprazine blocked amphetamine-induced hyperactivity (ED₅₀ = 0.12 mg/kg) and conditioned avoidance response (CAR) (ED₅₀ = 0.84 mg/kg) in rats, and inhibited the locomotor-stimulating effects of the noncompetitive NMDA antagonists MK-801 (ED₅₀ = 0.049 mg/kg) and phencyclidine (ED₅₀ = 0.09 mg/kg) in mice and rats, respectively. It reduced novelty-induced motor activity of mice (ED₅₀ = 0.11 mg/kg) and rats (ED₅₀ = 0.18 mg/kg) with a maximal effect of 70% in both species. Cariprazine produced no catalepsy in rats at up to 100-fold dose of its CAR inhibitory ED₅₀ value. Cariprazine 0.02-0.08 mg/kg significantly improved the learning performance of scopolamine-treated rats in a water-labyrinth learning paradigm. Though risperidone, olanzapine, and aripiprazole showed antipsychotic-like activity in many of these assays, they were less active against phencyclidine and more cataleptogenic than cariprazine, and had no significant effect in the learning task. The distinct in vivo profile of cariprazine may be due to its higher affinity and in vivo binding to D₃ receptors versus currently marketed typical and atypical antipsychotics.     

Anti-Analgesic Effect of the Mu/Delta Opioid Receptor Heteromer Revealed by Ligand-Biased Antagonism

Delta (DOR) and mu opioid receptors (MOR) can complex as heteromers, conferring functional properties in agonist binding, signaling and trafficking that can differ markedly from their homomeric counterparts. Because of these differences, DOR/MOR heteromers may be a novel therapeutic target in the treatment of pain. However, there are currently no ligands selective for DOR/MOR heteromers, and, consequently, their role in nociception remains unknown. In this study, we used a pharmacological opioid cocktail that selectively activates and stabilizes the DOR/MOR heteromer at the cell surface by blocking its endocytosis to assess its role in antinociception. We found that mice treated chronically with this drug cocktail showed a significant right shift in the ED₅₀ for opioid-mediated analgesia, while mice treated with a drug that promotes degradation of the heteromer did not. Furthermore, promoting degradation of the DOR/MOR heteromer after the right shift in the ED₅₀ had occurred, or blocking signal transduction from the stabilized DOR/MOR heteromer, shifted the ED₅₀ for analgesia back to the left. Taken together, these data suggest an anti-analgesic role for the DOR/MOR heteromer in pain. In conclusion, antagonists selective for DOR/MOR heteromer could provide an avenue for alleviating reduced analgesic response during chronic pain treatment.     

Comparison of three subcutaneous modes of prostaglandin F2α administration for pregnancy termination in the hamster

Dose response relationships for pregnancy termination in hamsters following administration of prostaglandin F_2α (PGF_2α by three subcutaneous methods were determined in 526 hamsters. The median effective dose (ED₅₀) for PGF_2α given as a single subcutaneous injection in 500 μl of saline was 22.2 μg. Administration of the prostaglandin with an Alzet® osmotic minipump (subcutaneous insertion for 24 hours) required 1.35 times more PGF_2α (ED₅₀ = 30.0 μg). The least effective method of prenancy termination in the hamster involved administration of PGF_2α by a single subcutaneous injection in 20.4 μl of saline (the same volume delivered by the minipump in 24 hours); the ED₅₀ for this method of administration was 41.3 μg of PGF_2α.