Biomedical subjects

R B Raffa

Publications and source records attributed to R B Raffa.

At least 55 records · Page 3Linked to original sources

PubMed1998

Tramadol and its enantiomers differentially suppress c-fos-like immunoreactivity in rat brain and spinal cord following acute noxious stimulus.

Tramadol hydrochloride, (1RS,2RS)-2-[(dimethylamino)methyl]-1-(3-methoxyphenyl)-cyclohexanol hydrochloride, is an orally-active, centrally-acting analgesic with a putative dual mechanism of action, including an opioid and non-opioid component. The analgesic properties of tramadol and the possible co-existence of dual mechanisms has been postulated to be due to complementary and interactive pharmacologies of its enantiomers. We examined the ability of tramadol, its enantiomers, and morphine as reference to suppress c-fos-like immunoreactivity (c-fos-ir) in rat spinal cord and brain regions following a noxious stimulus (i.p. administration of 3.5% acetic acid). c-fos-ir was measured by immunocytochemistry and the stained cells in each region were counted 2 h after the acetic-acid injection (2:25 h after tramadol or morphine). Equi-analgesic doses of s.c. morphine (10 mg/kg) or tramadol (30 mg/kg) significantly suppressed c-fos-ir in all areas examined, except dorsal central gray of the spinal cord. The enantiomers of tramadol had distinctive patterns of suppression, neither one suppressed c-fos-ir in all of the regions, and hence neither one alone accounted for the suppression produced by the racemate. These findings support differential and complementary effects of tramadol enantiomers in sub-populations of spinal and supraspinal nociceptive neurons, consistent with the proposed antinociceptive interaction between the enantiomers.

Journal Article↗

PubMed1998

Orphanin-FQ/nociceptin: lack of anti nociceptive, hyperalgesic or allodynic effects in acute thermal or mechanical tests following intracerebroventricular or intrathecal administration to mice or rats.

A recent review calls attention to the discrepant results resulting from studies that have examined the nociceptive or antinociceptive properties of orphanin-FQ/nociceptin (Phe-Gly-Gly-Phe-Thr-Gly-Ala-Arg-Lys-Ser-Ala-ArgLys-Leu-Ala-Asn-Gln; OFQ/N), the heptadecapeptide isolated from rat (nociceptin) and pig (orphanin FQ) brain that binds with high affinity to the opioid 'orphan' receptor (a seven transmembrane protein with sequence homology to opioid receptors), but exhibits only low affinity binding with conventional opioid ligands. Some of the discrepancy might result from differences in species, test, route of administration or time-course. We undertook a comprehensive examination of the effects of spinal (i.t.) or supraspinal (i.c.v.) administration of OFQ/N in mice and rats. Mice treated with OFQ/N either i.t. or i.c.v. demonstrated no significant nociceptive effect in the hot plate, warm-water or radiant heat tail-flick tests (except for the highest and most sedative dose of 10 nmol i.c.v. in the mouse warm-water tail-flick test). Pretreatment with the opioid antagonist naloxone or with peptidase inhibitors did not enhance the nociceptive effects of OFQ/N peptide in the warm-water tail-flick test. The motor activity in mice administered OFQ/N i.c.v. decreased significantly compared to controls. Rats administered i.c.v. or i.t. OFQ/N displayed no significant difference from vehicle-treated animals in similar noxious stimulus tests and OFQ/N-treated rats did not exhibit allodynia in a paw-withdrawal test. Overall, OFQ/N was ineffective in significantly altering response to noxious stimuli, regardless of whether the peptide was given at supraspinal or spinal sites in mice or in rats. In addition, i.c.v. or i.t. application of antisense or mismatch ODN to the orphan receptor did not modify tail-flick latency in either mice or rats, arguing against a tonic nociceptive tone mediated via the OFQ/N receptor.

Journal Article↗

PubMed1998

Neuropharmacology of St. John's Wort (Hypericum).

OBJECTIVE: To review preclinical information related to possible antidepressant mechanism(s) of action of St. John's wort in order to address the issue of whether its purported clinical effectiveness has a rational pharmacologic basis. DATA SOURCES: Primary and review articles were identified by a MEDLINE search (1966-January 1998) and through secondary sources. Many of the original German articles had English abstracts, but where necessary, German articles were translated into English. The results of a new screen of hypericin activity at receptor and uptake sites are summarized. STUDY SELECTION AND DATA EXTRACTION: All of the articles identified from the data sources were evaluated and all information deemed relevant was included in this review. DATA SYNTHESIS: The neuropharmacology of St. John's wort has been examined in only a few studies. A mechanism similar to that of the synthetic antidepressants, such as the selective serotonin-reuptake inhibitors or monoamine oxidase (MAO) inhibitors, might play a role, but other mechanisms are possible. CONCLUSIONS: Hypericum extracts have only weak activity in assays related to mechanisms of the synthetic antidepressants, that is, inhibition of MAO, catechol O-methyltransferase, or serotonin reuptake. It has been postulated that the clinical efficacy of St. John's wort could be attributable to the combined contribution of several mechanisms, each one too weak by itself to account for the overall effect. The recent demonstration of a significant affinity of hypericin for sigma receptors presents new possibilities for consideration.

Animals↗

PubMed1997

An examination of the relationship between mu-opioid antinociceptive efficacy and G-protein coupling using pertussis and cholera toxins.

The hypothesis that mu-opioid agonists having low antinociceptive efficacy might be more susceptible to interference with G-protein coupling than mu-opioid agonists having higher antinociceptive efficacy was tested. Supraspinal antinociceptive efficacy for the three mu-opioid agonists morphine, [D-Ala2, NMePhe4, Gly5-ol]-enkephalin (DAMGO) and sufentanil in the mouse 55 degrees C warm-water tail-flick test was evaluated 18-24 h after intracerebroventricular (i.c.v.) administration of beta-funaltrexamine (beta-FNA). The beta-FNA pretreatment (0.2-2.0 nmol) attenuated antinociception in the order morphine > DAMGO > sufentanil, consistent with previous reports of their relative antinociceptive efficacy. The association of efficacy with G-protein coupling was then assessed by determining sensitivity to i.c.v. (0.1-3.0 micrograms) pertussis toxin (PTX) or cholera toxin (CTX). The effect of PTX on equiantinociceptive doses was in the inverse order of agonist efficacy. CTX augmented sufentanil-induced antinociception. Morphine- and DAMGO-induced antinociception were unaffected by CTX. These data suggest that: (i) highly efficacious mu agonists (viz., sufentanil) couple more efficiently to PTX-sensitive inhibitory Gi-proteins than do agonists of lower efficacy (viz., morphine, DAMGO) and (ii) highly efficacious mu agonists have greater capacity to utilize CTX-sensitive stimulatory Gs-proteins than do mu-agonists with lower efficacy.

Analgesics↗

PubMed1997

Efficient designs for studying synergistic drug combinations.

Distinguishing between pharmacologically additive and synergistic drug combinations requires experimental designs and statistical analyses that often require appreciable numbers of animals and much experimenter time. The current study employed a design in which individual dose-effect data from each drug were translated into theoretically additive total dose combinations, in a fixed drug proportion, in order to produce a composite additive dose-effect relation that could be compared with that of an actual mixture having the same proportion. Results from this approach, using a combination of intrathecal doses of morphine and clonidine, were virtually identical to those using isobolographic analysis of the same data set. Both analyses showed significant synergism for this combination and, in each method, it was not necessary to constrain the drug regression lines to parallelism. In contrast to the isobole approach, the use of the composite additive dose-effect relation also allows observation of the interaction over a range of effects while reducing the size of the data sets needed.

Analgesia↗

PubMed1996

Tramadol, M1 metabolite and enantiomer affinities for cloned human opioid receptors expressed in transfected HN9.10 neuroblastoma cells.

Tramadol hydrochloride is a centrally acting synthetic analgesic in widespread clinical use. Despite different degrees of opioid-like characteristics in preclinical tests, it is characterized by lack of full naloxone reversibility or naloxone-precipitated withdrawal in humans. To investigate this apparent discrepancy, the present study measured the affinity of tramadol (and its enantiomers) and an active O-desmethyl metabolite (M1) (and its enantiomers) to cloned human opioid receptors of the mu, delta and kappa type stably expressed in HN9.10 neuroblastoma cells. At mu sites, the Ki values for tramadol, its (+) and (-) enantiomers, M1, and its (+) and (-) enantiomers were 17000, 15700, 28800, 3190, 153 and 9680 nM, respectively, compared to 7.1 nM for morphine. These results are consistent with the suggestion of a non-opioid contribution to the clinical profile of tramadol.

Brain Neoplasms↗

PubMed1996

A novel approach to the pharmacology of analgesics.

To date in the United States when a patient has presented with a complaint of pain requiring some form of pharmacologic relief, the physician has had the choice of two broad classes of drugs: peripherally acting (i.e., NSAID) or centrally acting (i.e., opioid) analgesics. The antidepressant monoamine reuptake inhibitors, particularly when combined with an opioid analgesic, have also proven efficacious in treating certain types of pain conditions. A new approach, available for almost 20 years in Europe and recently approved for use in the United States, is the centrally acting synthetic analgesic tramadol HCI. Preclinical evidence suggests that tramadol produces its antinociceptive effect in animals and analgesic effect in humans through a complementary dual mechanism of action. One mechanism relates to its weak affinity for mu-opioid receptors (6,000-fold less than morphine, 100-fold less than d-propoxyphene, 10-fold less than codeine, and equivalent to dextromethorphan). A metabolite (O-desmethyltramadol; M1) binds to opioid receptors with a greater affinity than the parent compound and could contribute to this component. However, in most animal tests and human clinical trials, the analgesic effect of tramadol is only partially blocked by the opioid antagonist naloxone, suggesting an important nonopioid mechanism. This nonopioid mechanism possibly relates to an increase in central neuronal synaptic levels of two neurotransmitters, 5-hydroxytryptamine (5-HT; serotonin) and norepinephrine. The opioid and nonopioid mechanisms appear to combine in a supra-additive manner in several tests of antinociception, but only in an additive or even counteracting manner in measures of adverse-effect liability. In sum, the apparent dual mechanism of action of tramadol suggests a possible new approach to pain relief.

Analgesics, Opioid↗

PubMed1996

Testing for synergism over a range of fixed ratio drug combinations: replacing the isobologram.

An isobologram is a Cartesian plot of pairs of doses that, in combination, yield a specified level of effect. It is a convenient and presently popular way of graphically displaying results of drug-combination and similar studies, because paired values of experimental points that fall below or above the line connecting the axial points (usually ED50 values) denote supra- and sub-additive combinations, respectively. However, an isobologram does not fulfill the criteria for standard least squares regression analysis. It is thus less useful for addressing questions related to the range of combination ratios over which synergy occurs. We describe herein a substitute for the isobologram in which log(total-dose) is plotted against the proportion of a component in a combination. One advantage is that a nonlinear curve-fitting procedure and determination of the confidence interval of a single parameter allow the determination of departure from additivity over a range of fixed proportion mixtures. An example is given of the combination of two analgesics (acetaminophen and tramadol hydrochloride). Another advantage of the new method is the reduction of animal use.

Acetaminophen↗

PubMed1996

A Gi2alpha antisense oligonucleotide differentiates morphine antinociception, constipation and acute dependence in mice.

In the same mice in which the intracerebroventricular (i.c.v.) administration of antisense oligodeoxyribonucleotide (oligo) directed against the Gi2alpha (but not Gi1alpha, Gi3alpha or G(s)alpha) G-protein subunits attenuated i.c.v. morphine-induced antinociception in the tail-flick test, none of the oligos altered naloxone-precipitated jumping (acute dependence). Likewise, none of the oligos significantly altered morphine-induced constipation. Hence, i.c.v. morphine-induced antinociception might be preferentially mediated via transduction pathway(s) different from constipation or acute dependence, offering novel opportunities for drug discovery.

Analgesics, Opioid↗

PubMed1996

Alpha-subunit G-protein antisense oligodeoxynucleotide effects on supraspinal (i.c.v.) alpha2-adrenoceptor antinociception in mice.

An in vivo antisense strategy was used to examine the involvement of G-protein subunits in supraspinal (intracerebroventricular; i.c.v.) alpha2-adrenoceptor-mediated antinociception. Mice that were injected with 33-mer antisense oligodeoxyribonucleotides (6 nmol) or vehicle were tested (tail-flick) with an agonist (clonidine, guanfacine or BH-T 920) administered i.c.v. 18 - 24 h later. Gi3alpha antisense treatment attenuated BH-T 920 and clonidine-induced antinociception. Gi2alpha antisense produced differential effects on the three agonists. Gi1alpha and G(s)alpha antisense treatment had no significant effect. Together with the previous demonstration that i.c.v. mu-opioid antinociception is mediated via Gi2alpha, the present results suggest that different receptors may mediate antinociception via different G-protein subunits and, hence, that specific subunits might offer novel targets for drug discovery.

Adrenergic alpha-Agonists↗

PubMed1996

Lack of binding of acetaminophen to 5-HT receptor or uptake sites (or eleven other binding/uptake assays).

The mechanism of analgesic action of acetaminophen (paracetamol) remains unknown. However, a central component distinct from that of the NSAIDs (non-steroidal antiinflammatory drugs) seems likely. A recent report (NeuroReport 6:1546-1548, 1995) suggests the involvement of 5-HT3 receptors. In the present study, we measured the affinity of acetaminophen at 5-HT3, as well as 5-HT1A, 5-HT1B, 5-HT1D, 5-HT2, 5-HT2C, 5-HT4, 5-HT6, 5-HT7 and eleven other receptor sites and at serotonin and norepinephrine reuptake sites. At 10 microM, acetaminophen inhibited less than 10% specific radioligand binding at any site. These findings: (i) suggest that acetaminophen's effect on serotonergic pathways is indirect, and (ii) circumscribe acetaminophen's possible central analgesic mechanism(s).

Acetaminophen↗

PubMed1996

Could dual G-protein coupling explain [D-Met2]FMRFamide's mixed action in vivo?

In vitro studies have demonstrated that FMRFamide-related peptide receptors can be coupled to different G-proteins, mediating opposite stimulatory and inhibitory effects. The present study tested whether this duality might extend to effects in vivo. Antinociception in mice of ICV [D-Met2]FMRFamide, which produced agonist [ED50 = 36.3 micrograms (61.6 nmol)] and antagonist [ID50 = 0.72 microgram (1.22 nmol)] actions, was attenuated by 24-h pretreatment with ICV pertussis toxin (ID50 = 0.55 microgram) or cholera toxin (ID50 = 0.09 microgram), suggesting that [D-Met2]FMRFamide in vivo effects might also be explained by dual Gi/Gs, coupling.

Amino Acid Sequence↗

PubMed1996

Endothelin-induced nociception in mice: mediation by ETA and ETB receptors.

Endothelins (ET-1, ET-2 or ET-3) or endothelin precursors (big-ET-1[1-38], big-ET-2[1-37] or big-ET-3[1-41]) injected i.p. in mice have previously been shown to elicit a characteristic nociceptive behavioral response. In this study, we investigated the endothelin receptor type (ETA or ETB) that mediates this behavioral response. Mice were injected i.p. with ET-1, ET-2, ET-3, big-ET-1[1-38], big-ET-2[1-37], big-ET-3[1-41], sarafotoxin S6a, sarafotoxin S6b, sarafotoxin S6c, ET-1 with Ala substitutions for Cys3 and Cys11 or His-Leu-Asp-Ile-Ile-Trp, and quantal dose-response curves were obtained for each of the compounds (except the latter). Co-administration of enzyme inhibitors with the big-endothelins was used to establish the requisite conversion to endothelins and big-ET-1[22-38], big-ET-2[22-37] and ET-3[22-41] amide, and the ETA-selective antagonist cyclo[-D-Asp-Pro-D-Val-Leu-D-Trp-] was used to determine receptor specificity. The ED50 values were 2.9, 3.3 and 23.9 micrograms/kg i.p. for ET-1, ET-2 and ET-3, respectively, 0.6, 0.6 and 13.1 micrograms/kg i.p. for sarafotoxin S6a, sarafotoxin S6b and sarafotoxin S6c, respectively, and 5.3 micrograms/kg i.p. for ET-1 with Ala substitutions for Cys3 and Cys11. Big-ET-1[22-38], big-ET-2[22-37], big-ET-3[22-41] amide and ET-C produced less than 25% effect up to 2000 micrograms/kg. The big-ET-1-induced effects were blocked by the enzyme inhibitors phosphoramidon and thiorphan (ID50 = 0.9 mg/kg) but not by ubenimex (bestatin), captopril or perindopril. Cyclo[-D-Asp-Pro-D-Val-Leu-D-Trp-] blocked ET-1- and ET-2-induced effects but not ET-3-, ACh- or phenyl-p-quinone-induced effects. These results suggest that endothelin-induced nociceptive behavioral response in mice can be mediated via both ET receptor types, ETA and ETB. Further, the ET-1 carboxy-terminal hexapeptide is insufficient to produce the effect, and the Cys3-Cys11 disulfide bridge of ET-1 is not required.

Animals↗

PubMed1996

Characterization of endothelin-induced nociception in mice: evidence for a mechanistically distinct analgesic model.

The behavioral response elicited in mice by an i.p. injection of endothelin-1 (ET-1) (0.1 mg/kg) was differentiated from that elicited by standard agents such as acetylcholine (ACh) (5.5 mg/kg) or phenyl-p-quinone (PpQ) (1.25 mg/kg). First, there was lack of two-way "cross-tolerance' between test paradigms. That is, at equieffective doses, a 60-min prior i.p. injection of ET-1 blocked the behavioral response to a subsequent i.p. injection of ET-1 or PpQ, but not of ACh, whereas a 60-min prior injection of ACh or of PpQ had no effect on a subsequent i.p. injection of ACh, PpQ or ET-1. Second, differential antagonism of ET-1-, ACh- or PpQ-induced responses was observed in an examination of 36 test compounds. For example, cyclo-oxygenase inhibitors such as indomethacin and ibuprofen did not block the ET-1-induced response at > 10 times the doses that blocked ACh- or PpQ-induced responses, whereas other compounds (such as certain benzodiazepines) inhibited ET-1-induced, but not ACh- or PpQ-induced, responses. These findings suggest that ET-1 produces a novel nociceptive stimulus, mechanistically distinct from ACh and PpQ. Hence, the ET-1-induced behavioral response in mice serves as a rapid and convenient measure of in vivo endothelin activity. In addition, this test might be a model for clinical pains not adequately treated by present analgesic agents or adequately tested by preclinical antinociceptive screens using ACh or PpQ. As such, it is a potentially valuable model for the identification of novel analgesic and other agents.

Acetylcholine↗

PubMed1996

Unexpected antinociceptive effect of the N-oxide (RWJ 38705) of tramadol hydrochloride.

N-Oxides of centrally acting analgesics generally have minimal analgesic activity. However, the N-oxide of tramadol produced dose-related, long-lasting antinociception in the mouse abdominal irritant, 48 degrees C hot-plate, 55 degrees C hot-plate, and tail-flick tests (ED50 = 15.5, 84.7, 316.4 and 138.2 mg/kg, p.o., respectively). Tramadol N-oxide (T-N-O) (RWJ 38705) was also antinociceptive in the 51 degrees C hot-plate test in male (ED50 = 63.2 mg/kg, i.p.) and female (ED50 = 39.9 mg/kg, i.p.) rats. A characteristic feature of T-N-O was an extended duration of action in these tests (4-5 h). T-N-O had negligible affinity for opioid mu (Ki = 38.5 microM) delta. or kappa receptors (Ki > 100 microM) and, in contrast to tramadol, was essentially devoid of norepinephrine or serotonin neuronal reuptake inhibitory activity (Ki > 100 microM). However, T-N-O displayed tramadol-like characteristics in vivo. There were also significant amounts of tramadol in plasma after T-N-O administration, and the levels resulting from equal oral doses of T-N-O and tramadol were the same, suggesting that the conversion of T-N-O to tramadol was rapid and essentially quantitative. T-N-O was not readily metabolized to tramadol in rat hepatic S9 fraction (< 2%), implying that the conversion might occur in the gastrointestinal tract. Taken together, the results suggest that T-N-O acts as a prodrug for tramadol. T-N-O could offer the clinical benefits of an extended duration of action and a "blunted" plasma concentration spike, possibly leading to an enhanced side-effect profile.

Analgesics↗

PubMed1995

Antinociceptive defect of beige-J mice reversed by i.c.v. IP3 or myo-inositol.

The C57BL/6J-bgJ/bgJ (beige-J) mutation imparts a blunted response to intracerebroventricular (i.c.v.) morphine in the tail-flick test, without altered micro-opioid receptor number or morphine affinity. We now report that co-administration of IP3 (36.1 nmol) restored morphine responsiveness of beige-J mice to essentially that of normal littermates (bg+/bg-; ED50 = 3.9 and 3.5 nmol, respectively). IP3 had no effect on morphine-induced antinociception in control animals. Neither myoinositol at 36.1 nmol nor IP6 at the highest testable dose (4.5 nmol) had a similar effect. Myo-inositol at 5.5 mumol restored beige-J responsiveness to that of littermates. These findings implicate some component of the phosphoinositide cycle in the antinociceptive defect of beige-J mice.

Analgesics↗

PubMed1995

The 'glibenclamide-shift' of centrally-acting antinociceptive agents in mice.

Morphine-induced antinociception is antagonized by the K(+)-channel blocker glibenclamide (glyburide; Glib), implicating ATP-sensitive (KATP) K+ channels in the analgesic effect of opioids. The present study examined the generality of this conclusion by measuring the effect of Glib on supraspinal (intracerebroventricular; i.c.v.) antinociception produced by representative mu-opioids and the non-opioids pilocarpine and two alpha 2-adrenoceptor agonists (clonidine and tizanidine) using the mouse tail-flick test. Concurrent administration of Glib (40 micrograms, i.c.v.) produced a significant rightward shift of the dose-response curve of morphine, levorphanol, methadone, pilocarpine, clonidine and tizanidine; a modest, but not statistically significant, rightward shift of the dose-response curves of the mu-selective peptides DAMGO ([D-Ala2,N-Me-Phe4,Gly-ol5]-enkephalin) and PL017 ([N-Me-Phe3,D-Pro4]-morphiceptin); and no shift of the dose-response curves of alfentanil, carfentanil, fentanyl, sufentanil, or beta-endorphin. Glib produced a leftward shift of the dose-response curve of etorphine. These data support the involvement of KATP-type K+ channels in mediation of supraspinal antinociception, differentiate Glib-sensitive and Glib-insensitive opioid agonists, and reveal fundamental differences among antinociceptive agents in the extent of demonstrable utilization of this transduction pathway.

Analgesics↗

PubMed1995

Alpha 2-adrenoceptors vs. imidazoline receptors: implications for alpha 2-mediated analgesia and other non-cardiovascular therapeutic uses.

The multiple clinical actions of clonidine have historically been linked to the same receptor (alpha 2-adrenoceptor) due to the belief that clonidine was a selective alpha 2-agonist. However, it is now recognized that clonidine binds with a similar affinity to alpha 2-adrenoceptors and to non-adrenergic imidazoline receptors. These two pharmacological targets (and subtypes of each alpha 2 and imidazoline receptors) provide the basis for a possible separation of cardiovascular and other targeted effects, such as analgesia. Consequently, the design of selective alpha 2-adrenoceptor (subtype) agonists as analgesics devoid of the cardiovascular effects associated with clonidine appears to be a rational approach to novel therapeutic agents. The present review focuses on alpha 2-adrenoceptor subtype/imidazoline diversity as a target for analgesic (and other CNS) drug discovery.

Adrenergic alpha-2 Receptor Agonists↗