RESUMEN
A diverse range of opioids and enantiomers were examined for their ability to displace binding at the [(3)H] MK-801-labelled site of the N-methyl-D-aspartate (NMDA) receptor in rat forebrain, displacement which is equitable with non-competitive NMDA receptor antagonist activity. Surprisingly, d-morphine, but not natural l-morphine, has low micromolar affinity for the site, suggesting clinical potential for racemic dl-morphine in the treatment of neuropathic pain with reduced development of tolerance. The opioid mu-receptor agonists: levorphanol, d- and dl-methadone, displayed similar properties. With the exception of the case of d-morphine, the structural requirements for affinity correspond closely with those previously found for the inhibitory effects of opioids on monoamine uptake.
Asunto(s)
Analgésicos Opioides/metabolismo , Maleato de Dizocilpina/metabolismo , Morfina/metabolismo , Prosencéfalo/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animales , Morfina/química , Norepinefrina/metabolismo , Dolor/tratamiento farmacológico , Dolor/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Serotonina/metabolismo , EstereoisomerismoRESUMEN
Human MPF (Lys-Lys-Gly-Glu) stimulates the proliferative response of human lymphocytes to the T-cell mitogen concanavalin A by 121-751% in the concentration range 10(-11)-10(-4) M; the peak effect is at 10(-8) M, lower or higher concentrations eliciting reduced responses, i.e. the dose-response curve is bell-shaped. Species specificity is high. Human MPF similarly stimulates rat lymphocytes, but the peak effect is seen at a 100-fold higher dose (10(-6) M). Rat MPF (Lys-Lys-Gly-Gln) has a peak effect at 10(-6) M with human lymphocytes, but the peak effect with rat lymphocytes is at a 1000-fold lower dose (10(-9) M). Truncated forms of the MPFs (Gly-Glu, Gly-Gln, Gly, Glu, Gln) and opioid peptides (beta-endorphin, [Leu] and [Met]enkephalin) show insignificant or only weak stimulatory or inhibitory effects. These results suggest that MPF acts via specific non-opioid receptors located on lymphocytes and that endogenously released MPF may have an important role in the functioning of the immune system.
Asunto(s)
Endorfinas/farmacología , Activación de Linfocitos/efectos de los fármacos , Oligopéptidos/farmacología , Fragmentos de Péptidos/farmacología , Animales , División Celular/efectos de los fármacos , División Celular/inmunología , Concanavalina A/farmacología , Dipéptidos/farmacología , Relación Dosis-Respuesta Inmunológica , Encefalina Leucina/farmacología , Encefalina Metionina/farmacología , Ácido Glutámico/farmacología , Glicina/farmacología , Humanos , Mitógenos/farmacología , Ratas , Especificidad de la Especie , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , betaendorfina/farmacologíaAsunto(s)
Analgésicos Opioides/uso terapéutico , Metadona/uso terapéutico , Morfina/uso terapéutico , Dolor Intratable/tratamiento farmacológico , Analgésicos Opioides/administración & dosificación , Analgésicos Opioides/efectos adversos , Humanos , Metadona/administración & dosificación , Metadona/efectos adversos , Morfina/administración & dosificación , Morfina/efectos adversosRESUMEN
MPF is a tetrapeptide (structure Lys-Lys-Gly-Glu) that elicits a variety of neurotrophic effects in vivo consistent with a role in neuronal regeneration. In support of this role, we now show that MPF stimulates the proliferation of cultured astrocytes and neurite outgrowth from cultures of neocortical cholinergic and mesenchephalic dopaminergic neurons. The dose-response relationships are biphasic ("bell shaped"), maximal responses being obtained with 10(-6) M concentrations of MPF. MPF and nerve growth factor seem to act on different receptors, because their effects on cholinergic neurons are synergistic.
Asunto(s)
Sistema Nervioso Central/citología , Sistema Nervioso Central/efectos de los fármacos , Endorfinas/farmacología , Fragmentos de Péptidos/farmacología , Hormona Adrenocorticotrópica/metabolismo , Animales , Astrocitos/citología , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , División Celular/efectos de los fármacos , Células Cultivadas , Sistema Nervioso Central/metabolismo , Corteza Cerebral/citología , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Citarabina/farmacología , Dopamina/metabolismo , Relación Dosis-Respuesta a Droga , Endorfinas/administración & dosificación , Humanos , Mesencéfalo/citología , Mesencéfalo/efectos de los fármacos , Mesencéfalo/metabolismo , Modelos Neurológicos , Factores de Crecimiento Nervioso/farmacología , Neuronas/citología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Fragmentos de Péptidos/administración & dosificación , RatasRESUMEN
We have previously shown that a metabolically stable analogue of MPF, the C-terminal tetrapeptide of human beta-endorphin of structure Lys-Lys-Gly-Glu, reduces the turning behaviour of rats with unilateral lesions of their nigro-striatal pathways. Transmission electron microscopy (TEM) has now revealed that this effect is related to reversal of the mitochondrial damage to substantia nigra (SN) neurones induced by the lesion. The results are consistent with the concept that an inherited defect in components of the mitochondrial enzyme system is the initial step in the genesis of Parkinson's disease (PD). They also, in conjunction with known neurotropic properties of MPF, and our unpublished finding of high concentrations of an MPF-like peptide in human basal ganglia, suggest that MPF may have physiological significance in the development and regeneration of the human CNS.
Asunto(s)
Factor Promotor de Maduración/uso terapéutico , Mitocondrias/ultraestructura , Enfermedad de Parkinson Secundaria/tratamiento farmacológico , Secuencia de Aminoácidos , Animales , Masculino , Microscopía Electrónica , Mitocondrias/efectos de los fármacos , Datos de Secuencia Molecular , Neuronas/efectos de los fármacos , Neuronas/ultraestructura , Oxidopamina , Enfermedad de Parkinson Secundaria/patología , Ratas , Ratas Wistar , Sustancia Negra/efectos de los fármacos , Sustancia Negra/patología , Sustancia Negra/ultraestructuraRESUMEN
A method is described for the determination of sub-picomole amounts of Lys-Lys-Gly-Glu [the C-terminal tetrapeptide sequence of human beta-endorphin, referred to as melanotropin potentiating factor (MPF)], a putative neurotrophic agent. Attempts to raise antibodies to the peptide were not successful and we have therefore developed a method based on the fluorescence of its 9-fluorenylmethyl chloroformate (FMOC) derivative which provides a sensitivity comparable to that of radioimmunoassay. Standard solutions, cerebrospinal fluid or central nervous tissue extracts are first treated with FMOC-Cl. The resulting mixture of FMOC-peptides is then subjected to high-performance liquid chromatography (HPLC) and quantified using a fluorescence monitor. By this procedure, MPF and related peptides can be analysed from one sample in a single HPLC run. The method was also applied to determine the rate of release into a phosphate-buffered saline medium of a metabolically stable analogue of MPF from a slow-release formulation of the compound.
Asunto(s)
Dipéptidos/análisis , Secuencia de Aminoácidos , Animales , Cromatografía Líquida de Alta Presión , Preparaciones de Acción Retardada , Dipéptidos/administración & dosificación , Dipéptidos/líquido cefalorraquídeo , Endorfinas , Fluorenos , Humanos , Indicadores y Reactivos , Masculino , Espectrometría de Masas , Datos de Secuencia Molecular , Fragmentos de Péptidos , Ratas , Ratas Wistar , TemperaturaRESUMEN
A single intrastriatal injection of a slow release formulation of a metabolically-stable MPF analogue was given to rats with lesioned right nigrostriatal pathways. After 6 weeks the turning behaviour of the rats in response to D-amphetamine began to decline, and after 12 weeks the reduction was marked and consistent. The implication of our results in the use of intracerebral grafts in parkinsonian patients is discussed.
Asunto(s)
Enfermedad de Parkinson/tratamiento farmacológico , Fragmentos de Péptidos/administración & dosificación , betaendorfina/análogos & derivados , Secuencia de Aminoácidos , Animales , Preparaciones de Acción Retardada , Modelos Animales de Enfermedad , Masculino , Datos de Secuencia Molecular , Ratas , Ratas Wistar , betaendorfina/administración & dosificaciónRESUMEN
The major metabolites of morphine, morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G), possess significant pharmacological properties. Whilst both metabolites bind to mu-opioid binding sites, M6G is a potent agonist whereas M3G antagonizes some of the effects of morphine and M6G. An inter-species comparison of in vitro hepatic morphine processing was performed. The results showed that not all species were able to produce M6G whereas all those tested did produce M3G. Guinea-pig liver produced the greatest amounts of M6G and was therefore chosen as a model to study morphine glucuronidation in further detail. Inclusion of the detergent Brij 58 (0.33 mg/mg protein) and Mg2+ (15 mM) in the standard assay incubation gave optimal production of both M3G and M6G by guinea-pig liver homogenates. A number of metal ions were investigated for their ability to inhibit glucuronidation of morphine in both the 3- and 6-positions. Some metal ions, namely Cu+, Cu2+ and Cd2+, were able to inhibit the production of M3G without affecting glucuronidation at the 6-position. Taken together, these data provide further evidence for the existence of UDP-glucuronosyltransferase isoenzymes responsible for the metabolism of morphine. In addition these isoenzymes can be differentially modulated and therefore it is possible to alter the ratio of M3G:M6G formed during in vitro metabolic studies.
Asunto(s)
Glucuronosiltransferasa/antagonistas & inhibidores , Hígado/metabolismo , Metales/farmacología , Morfina/metabolismo , Animales , Cadmio/farmacología , Cloruro de Cadmio , Cloruros/farmacología , Cobre/farmacología , Cobayas , Iones , Cinética , Hígado/efectos de los fármacos , Derivados de la Morfina/metabolismoRESUMEN
CSF methionine and leucine enkephalins were measured by high performance liquid chromatography and radioimmunoassay in diabetic patients with painful neuropathy (n = 22) and painless neuropathy (n = 5), and non-diabetic subjects with low back pain (n = 11). Wide variations in CSF enkephalin levels were found and they were often below the limit of detection (less than 0.1 pmol/l) in the diabetic and non-diabetic groups. The origin of CSF enkephalins is unknown and CSF levels may not reflect tissue concentrations. In conclusion, CSF enkephalin levels are difficult to interpret and do not provide useful information on the function of enkephalinergic pathways.
Asunto(s)
Neuropatías Diabéticas/líquido cefalorraquídeo , Encefalinas/líquido cefalorraquídeo , Adulto , Anciano , Anciano de 80 o más Años , Cromatografía Líquida de Alta Presión , Femenino , Humanos , Dolor de la Región Lumbar/líquido cefalorraquídeo , Masculino , Persona de Mediana Edad , RadioinmunoensayoRESUMEN
The output of dopamine from cultures of rat adrenal medullary strips was increased by 71-120% when Lys-Lys-Gly-Glu (MPF), the C-terminal tetrapeptide sequence of human beta-endorphin, was added to the culture medium at 100 micrograms/ml concentration. Human beta-endorphin caused a 44% increase, but an N-terminal fragment of its molecule and somatotrophin caused no increase. Results with analogs of MPF show that small structural change of the C-terminal Glu residue causes complete loss of activity.
Asunto(s)
Médula Suprarrenal/metabolismo , Dipéptidos/farmacología , Dopamina/metabolismo , Endorfinas/farmacología , Médula Suprarrenal/efectos de los fármacos , Animales , Endorfinas/metabolismo , Técnicas In Vitro , Fragmentos de Péptidos , Ratas , Ratas EndogámicasRESUMEN
The ability of B-endorphin to initiate limb regeneration in hypophysectomised newts is confined to the human species of the peptide and is contained in its C-terminal tetrapeptide sequence, Lys-Lys-Gly-Glu (MPF). Results with fifteen MPF analogs show that: (a) small structural change at the C-terminal Glu residue destroys activity, (b) at the Gly position, change of -NH-CH2-CO by -NH-NH-CO- (Azgly) or -NH-CHMe-CO- (Ala) also destroys the activity, but methylation of the NH results in an analog (Sar) with good activity, (c) analogs in which the Lys residues are replaced by D-Lys, Nle or Orn may retain some activity, particularly when the second Lys is replaced by D-Lys, and (d) the activity is retained or increased by N-terminal acylation. By combining 'favourable' changes, an analog acetyl-Lys-D-Lys-Sar-Glu was devised which was 1.25 times more potent than MPF, and metabolically more stable.
Asunto(s)
Extremidades/fisiología , Oligopéptidos/farmacología , Regeneración/efectos de los fármacos , betaendorfina/farmacología , Secuencia de Aminoácidos , Animales , Relación Dosis-Respuesta a Droga , Hipofisectomía , Inyecciones Intraperitoneales , Datos de Secuencia Molecular , Hormonas Hipofisarias/administración & dosificación , SalamandridaeRESUMEN
Most work on the function of peptides in neurons attempts to establish their role in transmission. Evidence in support of an alternative role in the growth and development of neurons and other cell types is discussed, and applied specifically in speculation concerning the etiology of chronic pain. Opportunities arise in the design of new types of drugs.
Asunto(s)
Diseño de Fármacos , Neuropéptidos/farmacología , Animales , Enfermedad Crónica , Humanos , Fenómenos Fisiológicos del Sistema Nervioso , Neuropéptidos/uso terapéutico , Dolor/tratamiento farmacológico , Dolor/fisiopatologíaRESUMEN
The ability of newts and other urolele amphibians to regenerate an accidentally-removed or amputated limb is lost after hypophysectomy, but restored following intraperitoneal injection of beta endorphin, or the C-terminal tetrapeptide sequence of human beta endorphin, Lys-Lys-Gly-Glu (= MPF). The closely related tetrapeptide, Lys-Lys-Gly-Gln (the C-terminal sequence of pig, sheep, and camel beta endorphin), the dipeptide Gly-Glu, the enkephalins, and other N-terminal sequences of beta endorphin (eg gamma endorphin) do not elicit the effect.
Asunto(s)
Endorfinas/farmacología , Fragmentos de Péptidos/farmacología , Regeneración/efectos de los fármacos , Animales , Miembro Anterior , Hipofisectomía , Hormonas Estimuladoras de los Melanocitos/farmacología , Salamandridae , Relación Estructura-Actividad , betaendorfina , gamma-EndorfinaRESUMEN
Highly specific antisera have been raised to [Leu]enkephalin and applied in a radioimmunoassay that is sufficiently sensitive and precise for the measurement of enkephalin pentapeptides in body fluids. Extraction of CSF by Sep-pak cartridges, followed by reverse-phase HPLC to separate [Leu] and [Met]enkephalin prior to assay, has assessed their concentrations as 59-170 pmol/l and 0.5-30 pmol/l, respectively, in three patients with chronic pain. The reproducibility of the HPLC separation step was checked by adding to the sample a synthetic analogue of very low cross-reactivity in UV detectable quantities. The assay procedure described is generally applicable to the assessment of [Leu] and [Met]enkephalin concentrations in CSF.