PubMed Health⌕ Search

Biomedical subjects

B L Appadu

Publications and source records attributed to B L Appadu.

11 recordsLinked to original sources
PubMed2000

Interaction of local anaesthetics with recombinant mu, kappa, and delta-opioid receptors expressed in Chinese hamster ovary cells.

Local anaesthetics potentiate epidural or intrathecal opioid analgesia via a poorly defined mechanism. In this study, we have examined the interaction of local anaesthetics (lidocaine, bupivacaine and its optical isomers, tetracaine, procaine and prilocaine) with recombinant mu-, kappa-, and delta-opioid receptors expressed in Chinese hamster ovary cells (CHO-mu, kappa, and delta, respectively). Lidocaine produced a concentration-dependent displacement of radiolabelled opioid antagonist [3H]diprenorphine ([3H]DPN) binding with the following rank order of inhibitor constant (Ki): kappa (210 microM) > mu (552 microM) > delta (1810 microM). Procaine, prilocaine, tetracaine and bupivacaine also displaced [3H]DPN binding in CHO-mu with Ki values of 244, 204, 43 and 161 microM respectively. Lidocaine produced a concentration-dependent and naloxone-insensitive inhibition of cAMP formation in all cell lines including untransfected cells. Concentration producing 50% inhibition of maximum was mu, 1.32 mM; kappa, 2.41 mM; delta, 1.27 mM; untransfected, 2.78 mM. When lidocaine (300 microM) was co-incubated with spiradoline (kappa-selective) and [D-Ala2, MePhe4, Gly(ol)5] enkephalin (DAMGO mu-selective) in CHO-kappa and mu cells we did not observe an additive interaction for cAMP formation. In contrast, there was an apparent inhibitory action of the combination at the kappa receptor. This study suggests that clinical concentrations of local anaesthetics interact with mu and kappa but not delta opioid receptors. As there was no synergism between local anaesthetics and opioids we suggest that the interaction of these agents in the clinical setting does not occur at the cellular level.

Anesthetics, Local↗
PubMed1999

Stereoselective interaction of ketamine with recombinant mu, kappa, and delta opioid receptors expressed in Chinese hamster ovary cells.

BACKGROUND: The authors examined the interaction of ketamine with recombinant mu, kappa, and delta opioid receptors and recombinant orphan opioid receptors expressed in Chinese hamster ovary cells (CHO-mu, CHO-kappa, CHO-delta, and CHO(ORL1), respectively). METHODS: CHO-mu, CHO-kappa, and CHO-delta membranes were incubated with the opioid receptor radioligand [3H]diprenorphine at room temperature. Ketamine (racemic, R(-) and S(+)) was included at concentrations covering the clinical range. CHO(ORL1) membranes were incubated with [125I]Tyr(14)nociceptin and racemic ketamine at room temperature. The effects of racemic ketamine and selective opioid receptor agonists (mu: [D-Ala2, MePhe4, Gly(ol)5] enkephalin (DAMGO); kappa: spiradoline or delta: [D-pen2, D-pen5] enkephalin (DPDPE)) on forskolin-stimulated cyclic adenosine monophosphate formation also were examined. Data are mean +/- SEM. RESULTS: Racemic ketamine increased the radioligand equilibrium dissociation constant for [3H]diprenorphine from 85+/-5 to 273+/-11, 91+/-6 to 154+/-16, and 372+/-15 to 855+/-42 pM in CHO-mu, CHO-kappa, and CHO-delta, respectively. The concentration of radioligand bound at saturation was unaffected. In CHO-mu and CHO-kappa cells, racemic ketamine did not slow the rate of naloxone-induced [3H]diprenorphine dissociation. Ketamine and its isomers also displaced [3H]diprenorphine binding to mu, kappa, and delta receptors in a dose-dependent manner, with pKi values for racemic ketamine of 4.38+/-0.02, 4.55+/-0.04, and 3.57+/-0.02, respectively. S(+)-ketamine was two to three times more potent than R(-)-ketamine at mu and kappa receptors. Racemic ketamine displaced [125I]Tyr(14)nociceptin with an estimated affinity constant of 0.5 mM. Racemic ketamine inhibited the formation of cyclic adenosine monophosphate (naloxone insensitive) in a dose-dependent manner (concentration producing 50% inhibition approximately 2 mM) in all cell lines, including untransfected CHO cells. Ketamine (100 microM) reversed DAMGO (mu) and spiradoline (kappa) inhibition of formation of cyclic adenosine monophosphate. CONCLUSIONS: Ketamine interacts stereoselectively with recombinant mu and kappa opioid receptors.

Animals↗
PubMed1998

Interaction of neuromuscular blocking drugs with recombinant human m1-m5 muscarinic receptors expressed in Chinese hamster ovary cells.

1. Neuromuscular blocking drugs (NMBD's) are known to produce cardiovascular side effects manifesting as brady/tachycardias. In this study we have examined the interaction of a range of steroidal NMBD's with recombinant human m1-m5 muscarinic receptors expressed in Chinese hamster ovary cells. Our main hypothesis is that NMBD's may interact with m2 (cardiac) muscarinic receptors. 2. All binding studies were performed with cell membranes prepared from CHO m1-m5 cells in 1 ml volumes of 20 mM HEPES, 1 mM MgCl2 at pH 7.4 for 1 h. Muscarinic receptors were labelled with [3H]-NMS and displacement studies were performed with pancuronium, vecuronium, pipecuronium, rocuronium and gallamine. In addition a range of muscarinic receptor subtype selective reference compounds were included. In order to determine the nature of any interaction the effects of pancuronium, rocuronium and vecuronium on methacholine inhibition of forskolin stimulated cyclic AMP formation in CHO m2 cells was examined. Cyclic AMP formation was assessed in whole cells using a radioreceptor assay. All data are mean +/- s.e.mean (n > or = 5). 3. The binding of [3H]-NMS was dose-dependent and saturable in all cells tested. Bmax and Kd values in m1-m5 cells were 2242+/-75, 165+/-13, 1877+/-33, 458+/-30, 127+/-2 fmol mg(-1) protein and 0.11+/-0.02, 0.15+/-0.01, 0.12+/-0.01, 0.12+/-0.01, 0.22+/-0.01 nM respectively. 4. The binding of [3H]-NMS was displaced dose dependently (pK50) by pirenzepine in CHO m1 membranes (7.97+/-0.04), methoctramine in CHO m2 membranes (8.55+/-0.1), 4-diphenylacetoxy-N-methyl piperidine methiodide (4-DAMP) in CHO m3 membranes (9.38+/-0.03), tropicamide in CHO m4 membranes (6.98+/-0.01). 4-DAMP, pirenzepine, tropicamide and methoctramine displaced [3H]NMS in CHO m5 membranes with pK50 values of 9.20+/-0.14, 6.59+/-0.04, 6.89+/-0.05 and 7.22+/-0.01 respectively. These data confirm homogenous subtype expression in CHO m1-m5 cells. 5. [3H]NMS binding was displaced dose-dependently (pK50) by pancuronium (m1, 6.43+/-0.12; m2, 7.68+/-0.02; m3, 6.53+/-0.06; m4, 6.56+/-0.03; m5, 5.79+/-0.10), vecuronium (m1, 6.14+/-0.04; m2, 6.90+/-0.05; m3, 6.17+/-0.04; m4, 7.31+/-0.02; m5, 6.20+/-0.07), pipecuronium (m1, 6.34+/-0.11; m2, 6.58+/-0.03; m3, 5.94+/-0.01; m4, 6.60+/-0.06; m5, 4.80+/-0.03), rocuronium (m1, 5.42+/-0.01; m2, 5.40+/-0.02; m3, 4.34+/-0.02; m4, 5.02+/-0.04; m5, 5.10+/-0.03) and gallamine (m1, 6.83+/-0.05; m2, 7.67+/-0.04; m3, 6.06+/-0.06; m4, 6.20+/-0.03; m5, 5.34+/-0.03). 6. Cyclic AMP formation was inhibited dose dependently by methacholine in CHO m2 cells pEC50 for control and pancuronium (300 nM) treated cells were 6.18+/-0.34 and 3.57+/-0.36 respectively. Methacholine dose-response curves in the absence and presence of rocuronium (1 microM) and vecuronium (1 microM) did not differ significantly. Pancuronium, vecuronium and rocuronium did not inhibit cyclic AMP formation alone indicating no agonist activity. 7. With the exception of rocuronium there was a significant interaction with m2 muscarinic receptors with all NMBD's at clinically achievable concentrations suggesting that the brady/tachycardias associated with these agents may result from an interaction with cardiac muscarinic receptors. Furthermore pancuronium at clinically achievable concentrations antagonised methacholine inhibition of cyclic AMP formation in CHO m2 cells further suggesting that the tachycardia produced by this agent results from muscarinic antagonism. The mechanism of the bradycardia produced by vecuronium is unclear.

Analysis of Variance↗
PubMed1997

Do local anaesthetics interact with dihydropyridine binding sites on neuronal L-type Ca2+ channels?

We have examined the interaction of procaine, prilocaine, lignocaine, bupivacaine, amylocaine and R(+) and S(-) ropivacaine with L-type voltage-sensitive Ca2+ channels in rat cerebrocortical membranes. Membranes were prepared in Tris HCl 50 mmol litre-1, pH 7.4, by homogenization and centrifugation. Binding assays were performed in 1-ml volumes of Tris HCl 50 mmol litre-1, pH 7.4, for 90 min at room temperature using approximately 200 micrograms of protein. Non-specific binding was defined in the presence of nifedipine 10(-5) mol litre-1, and bound and free radioactivity were separated by vacuum filtration. The effects of local anaesthetics were determined by displacement of [3H]PN200-110 (approximately 0.2 nmol litre-1), a radiolabelled 1,4- dihydropyridine (DHP) L-channel antagonist. The concentration of displacer producing 50% displacement was corrected for the competing mass of [3H]PN200-110 to yield the affinity constant, K50. All local anaesthetics displaced [3H]PN200-110 in a dose-dependent manner with a rank order potency of (K50, mmol litre-1) bupivacaine (0.48), amylocaine (0.74), lignocaine (1.09), prilocaine (2.06) and procaine (2.09). Ropivacaine enantiomers did not show stereo-selective displacement, with K50 values of 0.99 and 0.92 mmol litre-1 for R(+) and S(-) ropivacaine, respectively. There was a significant correlation between pK50 and p (octanol:buffer partition coefficient) (r2 = 0.872, P = 0.020), pK50 and p (local anaesthetic potency) (r2 = 0.816, P = 0.036), pK50 and p (relative conduction blocking potency) (r2 = 0.843, P = 0.028) and between pK50 and p (IC50 for inhibition of cardiac output) (r2 = 0.897, P = 0.015). These data suggest that DHP binding sites may be involved in both the mechanism of local anaesthesia and the cardiotoxicity of these agents.

Anesthetics, Local↗
PubMed1997

Anaesthetic potency of inhalation agents is independent of membrane microviscosity.

The decrease in membrane microviscosity of erythrocyte ghosts in the presence of clinically relevant concentrations of seven inhalation anaesthetic agents was studied using fluorescence polarization anisotropy of the membrane incorporated fluorescent probes 1,6-diphenyl-1,3,5-hexatriene and 1-[4-trimethylammoniumphenyl]-6-phenyl-1,3,5-hexatriene. All anaesthetic agents produced a dose-dependent decrease in anisotropy of both probes, indicating decreased membrane microviscosity. The reduction in anisotropy measured at the minimum alveolar concentration (ED50) for anaesthesia was related inversely to the anaesthetic potency of the agent and was directly proportional to the hypothetical concentration of agent in the membrane calculated from lipid-water partition coefficients. These findings do not support the hypothesis that volatile anaesthetic agents act by increasing membrane microviscosity of the bulk lipid bilayer to produce anaesthesia.

Anesthetics, Inhalation↗
PubMed1996

Postal survey on the long-term use of neuromuscular block in the intensive care.

OBJECTIVE: To assess the long-term use of neuromuscular blocking (NMB) agents in intensive care, especially with reference to the potential problems of the long-term use of NMB drugs in the intensive care unit (ICU). METHOD: A postal survey questionnaire was sent to 409 ICUs in Great Britain. RESULTS: Two hundred thirty-eight completed questionnaires were returned and analysed. Most ICUs were anaesthetist-led (85.8%) with only five ICUs being staffed by full-time intensivists. Facilitation of mechanical ventilation and increased intracranial pressure were the main indications for the prolonged use of neuromuscular blockade. Atracurium and vecuronium (83%) were administered most commonly by bolus alone (13.8%), bolus followed by continuous infusion (23.9%) or continuous infusion only (60.9%). The most frequently cited criteria for the use of either vecuronium or stracurium were their pharmacokinetics and haemodynamic stability. Neuromuscular block was most commonly monitored clinically (91.7%), with only 8.3% of the responders using a peripheral nerve stimulator. All responders indicated the concomitant use of sedatives (propofol/midazolam alone or in combination in 89.4% of responders) and/or opioids (morphine, fentanyl or alfentanil in 74.8% of respondents) with muscle relaxants. CONCLUSION: Most responders agreed that while neuromuscular block in the ICU population may provide advantages, it cannot be considered benign. Indeed, a great majority consider that NMB agents should be used only as a last option and -for as short a period as possible.

Analgesics, Opioid↗
PubMed1996

Interaction of i.v. anaesthetic agents with 5-HT3 receptors.

Using N1E-115 neuroblastoma cells as an experimental model, we have examined if four commonly used i.v. anaesthetic induction agents interact with 5-HT3 receptors. Specifically, we tested the hypothesis that the antiemetic effects of propofol may result from 5-HT3 receptor antagonism. Binding of tropisetron (a 5-HT3 selective reference compound), etomidate, ketamine, thiopentone and propofol to 5-HT3 receptors was assessed by measuring the displacement of [3H]BRL 43694 from whole N1E-115 cells. The rank order potency (Ki) was tropisetron (1.7 (SEM 0.2) nmol litre-1) >> etomidate (83.(4) mumol litre-1) > or = ketamine (97 (4) mumol litre-1) > thiopentone (177 (9) mumol litre-1) > propofol (819 (171) mumol litre-1). With the exception of thiopentone these effects were outside the clinical range and suggest that anaesthetic agents are unlikely to interact directly with 5-HT3 receptors, and that other mechanism(s) must underlie the antiemetic effects of propofol.

Anesthetics, Intravenous↗
PubMed1994

Studies on the interaction of steroidal neuromuscular blocking drugs with cardiac muscarinic receptors.

Interaction of the steroidal neuromuscular blocking drugs pancuronium, pipecuronium, rocuronium and vecuronium with cardiac muscarinic receptors in rat hearts was investigated in vitro by a tritiated N-methyl hyoscine binding assay. We showed an interaction with cardiac muscarinic receptors with a rank order of potency pancuronium > vecuronium > pipecuronium > rocuronium and demonstrated complex binding characteristics for pancuronium, vecuronium and rocuronium, with "Hill coefficient" of less than unity. We conclude that the haemodynamic differences seen during the use of these neuromuscular blocking drugs may be a result of their interactions with cardiac M2 muscarinic receptors.

Androstanols↗