The Endogenous Cannabinoid Anandamide Inhibits Cromakalim-Activated K+ Currents in Follicle-Enclosed Xenopus Oocytes

The effect of the endogenous cannabinoid anandamide on K+ currents activated by the ATP-sensitive potassium (KATP) channel opener cromakalim was investigated in follicle-enclosed Xenopus oocytes using the two-electrode voltage-clamp technique. Anandamide (1–90 μM) reversibly inhibited cromakalim-induced K+ currents, with an IC50 value of 8.1 ± 2 μM. Inhibition was noncompetitive and independent of membrane potential. Coapplication of anandamide with the cannabinoid type 1 (CB1) receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride (SR 141716A) (1 μM), the CB2 receptor antagonist N-[(1S)endo-1,3,3-trimethyl bicyclo heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528) (1 μM), or pertussis toxin (5 μg/ml) did not alter the inhibitory effect of anandamide, suggesting that known cannabinoid receptors are not involved in anandamide inhibition of K+ currents. Similarly, neither the amidohydrolase inhibitor phenylmethylsulfonyl fluoride (0.2 mM) nor the cyclooxygenase inhibitor indomethacin (5 μM) affected anandamide inhibition of K+ currents, suggesting that the effects of anandamide are not mediated by its metabolic products. In radioligand binding studies, anandamide inhibited the specific binding of the KATP ligand [3H]glibenclamide in the oocyte microsomal fractions, with an IC50 value of 6.3 ± 0.4 μM. Gonadotropin-induced oocyte maturation and the cromakalim-acceleration of progesterone-induced oocyte maturation were significantly inhibited in the presence of 10 μM anandamide. Collectively, these results indicate that cromakalim-activated K+ currents in follicular cells of Xenopus oocytes are modulated by anandamide via a cannabinoid receptor-independent mechanism and that the inhibition of these channels by anandamide alters the responsiveness of oocytes to gonadotropin and progesterone

Cholecystokinin B-type Receptors Mediate a G-Protein-Dependent Depolarizing Action of Sulphated Cholecystokinin Ocatapeptide (CCK-8s) on Rodent Neonatal Spinal Ventral Horn Neurons

Reports of cholecystokinin (CCK) binding and expression of CCK receptors in neonatal rodent spinal cord suggest that CCK may influence neuronal excitability. In patch-clamp recordings from 19/21 ventral horn motoneurons in neonatal (PN 5–12 days) rat spinal cord slices, we noted a slowly rising and prolonged membrane depolarization induced by bath-applied sulfated CCK octapeptide (CCK-8s; 1 μM), blockable by the CCKB receptor antagonist L-365,260 (1 μM). Responses to nonsulfated CCK-8 or CCK-4 were significantly weaker. Under voltage clamp (VH −65 mV), 22/24 motoneurons displayed a CCK-8s-induced tetrodotoxin-resistant inward current [peak: −136 ± 28 pA] with a similar time course, mediated via reduction in a potassium conductance. In 29/31 unidentified neurons, CCK-8s induced a significantly smaller inward current (peak: −42.8 ± 5.6 pA), and I-V plots revealed either membrane conductance decrease with net inward current reversal at 101.3 ± 4.4 mV (n = 16), membrane conductance increase with net current reversing at 36.1 ± 3.8 mV (n = 4), or parallel shift (n = 9). Intracellular GTP-γ-S significantly prolonged the effect of CCK-8s (n = 6), whereas GDP-β-S significantly reduced the CCK-8s response (n = 6). Peak inward currents were significantly reduced after 5-min perfusion with N-ethylmaleimide. In isolated neonatal mouse spinal cord preparations, CCK-8s (30–300 nM) increased the amplitude and discharge of spontaneous depolarizations recorded from lumbosacral ventral roots. These observations imply functional postsynaptic G-protein-coupled CCKB receptors are prevalent in neonatal rodent spinal cord

The Nonpsychoactive Cannabinoid Cannabidiol Inhibits 5-Hydroxytryptamine3A Receptor-Mediated Currents in Xenopus laevis Oocytes

The effect of the plant-derived nonpsychotropic cannabinoid, cannabidiol (CBD), on the function of hydroxytryptamine (5-HT)3A receptors expressed in Xenopus laevis oocytes was investigated using two-electrode voltage-clamp techniques. CBD reversibly inhibited 5-HT (1 μM)-evoked currents in a concentration-dependent manner (IC50 = 0.6 μM). CBD (1 μM) did not alter specific binding of the 5-HT3A antagonist [3H]3-(5-methyl-1H-imidazol-4-yl)-1-(1-methylindol-3-yl)propan-1-one (GR65630), in oocytes expressing 5-HT3A receptors. In the presence of 1 μM CBD, the maximal 5-HT-induced currents were also inhibited. The EC50 values were 1.2 and 1.4 μM, in the absence and presence of CBD, indicating that CBD acts as a noncompetitive antagonist of 5-HT3 receptors. Neither intracellular BAPTA injection nor pertussis toxin pretreatment (5 μg/ml) altered the CBD-evoked inhibition of 5-HT-induced currents. CBD inhibition was inversely correlated with 5-HT3A expression levels and mean 5-HT3 receptor current density. Pretreatment with actinomycin D, which inhibits protein transcription, decreased the mean 5-HT3 receptor current density and increased the magnitude of CBD inhibition. These data demonstrate that CBD is an allosteric inhibitor of 5-HT3 receptors expressed in X. laevis oocytes. They further suggest that allosteric inhibition of 5-HT3 receptors by CBD may contribute to its physiological roles in the modulation of nociception and emesis

Gender Differences in the Behavioral Responses to Cocaine and Amphetamine

When ovariectomized female rats receive estrogen, the response to the psychomotor stimulants amphetamine or cocaine is enhanced. Estrous cycle-dependent differences in amphetamine-stimulated behaviors and striatal dopamine release are also noted. Intact female rats exhibit a greater behavioral response to amphetamine on estrus than they do on other days of the cycle. Ovariectomy results in attenuation of amphetamine-induced behavior and the striatal dopamine response to amphetamine. Physiological doses of estrogen given to ovariectomized rats reinstate both of these responses to a level comparable to that in estrous females. Furthermore, a sex difference is noted, in that females tend to exhibit a greater behavioral response to the psychomotor stimulants, and estrogen enhances this sex difference. Repeated treatment with amphetamine or cocaine produces a progressive increase in behavioral responsiveness with subsequent drug administration, a process known as sensitization. In rodents, behavioral sensitization results in increases in both frequency and duration of psychomotor behaviors such as rotational behavior, stereotyped grooming, headbobs, and forelimb movements. Interestingly, females display greater sensitization of behaviors in response to psychomotor stimulants than do males. Previous research results are summarized, and new results are presented, demonstrating that estrogen selectively enhances components of behavior that exhibit sensitization in female rats. Results also indicate gender differences in sensitization independent of gonadal hormones, suggesting that the neural systems that undergo sensitization are sexually dimorphic.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/72269/1/j.1749-6632.2001.tb03564.x.pd

Dopamine receptor mRNA expression patterns by opioid peptide cells in the nucleus accumbens of the rat: A double in situ hybridization study

Colocalization of proenkephalin and prodynorphin mRNAs with each other as well as with D1, D2, and D3 dopamine receptor mRNAs was analyzed in the nucleus accumbens of the rat. Distinct combinations were detected in the rostral pole, core, and shell subdivisions of the nucleus accumbens. Proenkephalin and prodynorphin mRNAs were principally localized in separate cells in the core. All detectable prodynor cells in the core expressed D1 mRNA but not D2 mRNA. Conversely, approximately 95% of the proenkephalin-positive cells in this region expressed D2 mRNA but not D1 mRNA. This pattern was identical to that observed in the caudate putamen. In the rostral pole and the shell, embedded in a background of this “typical” colocalization pattern, clusters of cells expressing a distinct configuration were found. In these clusters, proenkephalin-positive cells expressed both prodynorphin and D1 mRNAs, but they did not express D2 mRNA. D3 and prodynorphin mRNAs were colocalized in “limbic” striatal areas, including the ventromedial caudate putamen, the rostral pole, and the medial shell. In contrast, D3 mRNA was not detected in any proenkephalin-positive cells. Together with the prodynorphin/D1 data, this suggests that a subset of prodynorphin cells expresses both D1 and D3 mRNAs. It is concluded that (1) clusters of cells that coexpress proenkephalin, prodynorphin, and D1 mRNAs overlap extensively with previously defined cytoarchitectural cell clusters in the nucleus accumbens and (2) a subset of the prodynorphin cells in the ventromedial caudate putamen and the nucleus accumbens contains both D1 and D3 mRNAs. © 1995 Wiley-Liss, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/50067/1/903610106_ftp.pd

Reference place conditioning procedure with cocaine: Increased sensitivity for measuring associatively motivated choice behavior in rats

Place conditioning is widely used to study the conditioned rewarding effects of drugs. In the standard version, one reward (cocaine) is compared to no reward (saline). A modified variant of this task, “reference-conditioning” procedure, compares two potentially rewarding stimuli (high versus low cocaine dose). There has been little research on the utility of this procedure. Experiment 1 used the standard protocol with saline administered before confinement to the reference compartment of a place-conditioning chamber. On alternating days, saline, 2.5, 5, 7.5, 10, or 20 mg/kg cocaine was administered before confinement to the opposite compartment. In Experiments 2 and 3, reference-compartment saline was replaced with 5 and 7.5 mg/kg cocaine, respectively. Relative to saline, 7.5–20 mg/kg cocaine had comparable conditioned rewarding effects (i.e., similar increase in time in paired compartment). When cocaine replaced saline, there was competition at doses lower than 7.5 mg/kg. Rats that received 7.5 versus 2.5 mg/kg spent similar time in each compartment, indicating competition. Competition was not seen with 5 versus 20 mg/kg; preference was for the 20 mg/kg compartment. Experiment 4 showed that the competition at 2.5 mg/kg was not due to reward sensitization—. The reference-conditioning procedure has increased sensitivity for measuring associatively-motivated choice behavior

Salvinorin A Regulates Dopamine Transporter Function Via A Kappa Opioid Receptor and ERK1/2-Dependent Mechanism

Salvinorin A (SalA), a selective κ-opioid receptor (KOR) agonist, produces dysphoria and pro-depressant like effects. These actions have been attributed to inhibition of striatal dopamine release. The dopamine transporter (DAT) regulates dopamine transmission via uptake of released neurotransmitter. KORs are apposed to DAT in dopamine nerve terminals suggesting an additional target by which SalA modulates dopamine transmission. SalA produced a concentration-dependent, nor-binaltorphimine (BNI)- and pertussis toxin-sensitive increase of ASP+ accumulation in EM4 cells coexpressing myc-KOR and YFP-DAT, using live cell imaging and the fluorescent monoamine transporter substrate, trans 4-(4-(dimethylamino)-styryl)-N-methylpyridinium) (ASP+). Other KOR agonists also increased DAT activity that was abolished by BNI pretreatment. While SalA increased DAT activity, SalA treatment decreased serotonin transporter (SERT) activity and had no effect on norepinephrine transporter (NET) activity. In striatum, SalA increased the Vmax for DAT mediated DA transport and DAT surface expression. SalA up-regulation of DAT function is mediated by KOR activation and the KOR-linked extracellular signal regulated kinase-½ (ERK1/2) pathway. Co-immunoprecipitation and BRET studies revealed that DAT and KOR exist in a complex. In live cells, DAT and KOR exhibited robust FRET signals under basal conditions. SalA exposure caused a rapid and significant increase of the FRET signal. This suggests that the formation of KOR and DAT complexes is promoted in response to KOR activation. Together, these data suggest that enhanced DA transport and decreased DA release resulting in decreased dopamine signaling may contribute to the dysphoric and pro-depressant like effects of SalA and other KOR agonists

Role of protein kinase C and μ-opioid receptor (MOPr) desensitization in tolerance to morphine in rat locus coeruleus neurons

In morphine tolerance a key question that remains to be answered is whether μ-opioid receptor (MOPr) desensitization contributes to morphine tolerance, and if so by what cellular mechanisms. Here we demonstrate that MOPr desensitization can be observed in single rat brainstem locus coeruleus (LC) neurons following either prolonged (> 4 h) exposure to morphine in vitro or following treatment of animals with morphine in vivo for 3 days. Analysis of receptor function by an operational model indicated that with either treatment morphine could induce a profound degree (70–80%) of loss of receptor function. Ongoing PKC activity in the MOPr-expressing neurons themselves, primarily by PKCα, was required to maintain morphine-induced MOPr desensitization, because exposure to PKC inhibitors for only the last 30–50 min of exposure to morphine reduced the MOPr desensitization that was induced both in vitro and in vivo. The presence of morphine was also required for maintenance of desensitization, as washout of morphine for > 2 h reversed MOPr desensitization. MOPr desensitization was homologous, as there was no change in α2-adrenoceptor or ORL1 receptor function. These results demonstrate that prolonged morphine treatment induces extensive homologous desensitization of MOPrs in mature neurons, that this desensitization has a significant PKC-dependent component and that this desensitization underlies the maintenance of morphine tolerance

Opioid receptors in GtoPdb v.2025.3

Opioid and opioid-like receptors are activated by a variety of endogenous peptides including [Met]enkephalin (met), [Leu]enkephalin (leu), β-endorphin (β-end), α-neodynorphin, dynorphin A (dynA), dynorphin B (dynB), big dynorphin (Big dyn), nociceptin/orphanin FQ (N/OFQ); endomorphin-1 and endomorphin-2 are also potential endogenous peptides. The Greek letter nomenclature for the opioid receptors, μ, δ and κ, is well established, and NC-IUPHAR considers this nomenclature appropriate, along with the symbols spelled out (mu, delta, and kappa), and the acronyms, MOP, DOP, and KOP [127, 103, 94]. However the acronyms MOR, DOR and KOR are still widely used in the literature. The human N/OFQ receptor, NOP, is considered \u27opioid-related\u27 rather than opioid because, while it exhibits a high degree of structural homology with the conventional opioid receptors [313, 94], it displays a distinct pharmacology. Currently there are numerous clinically used drugs, such as morphine and many other opioid analgesics, as well as antagonists such as naloxone. The majority of clinically used opiates are relatively selective μ agonists or partial agonists, though there are some μ/κ compounds, such as butorphanol, in clinical use. κ opioid agonists, such as the alkaloid nalfurafine and the peripherally acting peptide difelikefalin, are in clinical use for itch

Opioid receptors in GtoPdb v.2025.1

Opioid and opioid-like receptors are activated by a variety of endogenous peptides including [Met]enkephalin (met), [Leu]enkephalin (leu), β-endorphin (β-end), α-neodynorphin, dynorphin A (dynA), dynorphin B (dynB), big dynorphin (Big dyn), nociceptin/orphanin FQ (N/OFQ); endomorphin-1 and endomorphin-2 are also potential endogenous peptides. The Greek letter nomenclature for the opioid receptors, μ, δ and κ, is well established, and NC-IUPHAR considers this nomenclature appropriate, along with the symbols spelled out (mu, delta, and kappa), and the acronyms, MOP, DOP, and KOP [126, 103, 94]. However the acronyms MOR, DOR and KOR are still widely used in the literature. The human N/OFQ receptor, NOP, is considered \u27opioid-related\u27 rather than opioid because, while it exhibits a high degree of structural homology with the conventional opioid receptors [310], it displays a distinct pharmacology. Currently there are numerous clinically used drugs, such as morphine and many other opioid analgesics, as well as antagonists such as naloxone. The majority of clinically used opiates are relatively selective μ agonists or partial agonists, though there are some μ/κ compounds, such as butorphanol, in clinical use. κ opioid agonists, such as the alkaloid nalfurafine and the peripherally acting peptide difelikefalin, are in clinical use for itch