RESUMEN

Avy/agouti (Avy) mice have late onset obesity related to overexpression of agouti signaling protein (ASP) in the hypothalamus. As mahogany modulates the actions of ASP, we tested the transport of mahogany peptide across the blood-brain barrier (BBB). The brain uptake of mahogany peptide was significantly higher in young Avy mice, and it preceded the surge of fat mass quantified by nuclear magnetic resonance. The results suggest a role of accelerated BBB transport in the epigenetics of Avy mice.

Asunto(s)

Tejido Adiposo/metabolismo , Barrera Hematoencefálica/metabolismo , Proteínas de la Membrana/farmacocinética , Proteína de Señalización Agouti , Animales , Transporte Biológico/fisiología , Encéfalo/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Imagen por Resonancia Magnética , Ratones , Ratones Mutantes , Obesidad/metabolismo

RESUMEN

The blood-brain barrier (BBB) plays a crucial role in the regulation of body weight and feeding by peptides. This review summarizes the components of the BBB as well as the circumventricular organs (CVOs), the methods used for quantification of the passage of feeding peptides across the BBB, and the various ways by which these peptides can interact with the BBB.

Asunto(s)

Barrera Hematoencefálica/fisiología , Conducta Alimentaria/fisiología , Péptidos/fisiología , Animales , Transporte Biológico/fisiología , Humanos , Péptidos/metabolismo

RESUMEN

Endomorphins are endogenous opioid peptides with high affinity and selectivity for the mu-opioid receptor. In the present study, we examined the morphology of the endomorphin 2-like immunoreactive (EM2-LI) neurons in the hypothalamus at the light and electron microscopic levels. At the light microscopic level, EM2-LI neurons were found mostly distributed in the regions between the dorsomedial and ventromedial hypothalamic nuclei and the region near the third ventricle. At the electron microscopic level, EM2-LI perikarya could be divided into two groups. Type I perikarya contained relatively undeveloped endoplasmic reticulum and Golgi apparatus while type II perikarya contained well-developed rough-surfaced endoplasmic reticulum and Golgi apparatus. Both type I and type II neurons contained numerous EM2-LI dense-cored vesicles. Type II perikarya and dendrites received synapses and showed immunoreactivity in the endoplasmic reticulum and Golgi apparatus. EM2-LI axon terminals formed synapses with both immunonegative and immunopositive dendrites. In some cases, the axon terminals contained both immunonegative and immunopositive dense-cored vesicles. EM2-LI neurons often had synaptic relationships with neurons containing immunonegative dense-cored vesicles. Myelinated axon shafts containing EM2-LI were also found. This first demonstration of the ultrastructure and synaptic relationships of EM2-LI neurons in the hypothalamus provides morphological evidence that suggests (1) endomorphin 2-containing neurons modulate physiological function through synaptic relationships; (2) endomorphin 2 may coexist with other neurotransmitters in the same neurons; and (3) endomorphin 2-containing neurons could modulate other endomorphin 2-containing neurons as well as those containing other neurotransmitters.

Asunto(s)

Neuronas/metabolismo , Oligopéptidos/metabolismo , Sinapsis/ultraestructura , Animales , Hipotálamo Posterior/citología , Hipotálamo Posterior/metabolismo , Hipotálamo Posterior/ultraestructura , Inmunohistoquímica , Cuerpos de Inclusión/metabolismo , Masculino , Microscopía Inmunoelectrónica , Vías Nerviosas/metabolismo , Vías Nerviosas/ultraestructura , Neuronas/citología , Neuronas/ultraestructura , Ratas , Ratas Wistar , Sinapsis/metabolismo

RESUMEN

OBJECTIVE: Peripherally administered exendin-4 is in clinical trials for the treatment of diabetes mellitus and obesity. Since its effects on food intake are mediated centrally, we determined the degree and type of its blood-to-brain penetration of the mouse blood-brain barrier (BBB). MEASUREMENTS AND RESULTS: High-performance liquid chromatography showed that exendin-4 was stable in blood, with most of the injected peptide reaching the brain intact. Capillary depletion studies with washout showed that the injected exendin-4 reached brain parenchyma rather than being trapped in the endothelial cells composing the BBB. Multiple-time regression analysis showed that exendin-4 crossed the BBB directly at a fast rate. The rapid brain entry of exendin-4, helped by its high lipophilicity as demonstrated by the octanol/buffer partition coefficient, was not dependent upon circumventricular organs and was not affected by food deprivation for 24 h. The simultaneous i.v. injection of high doses of unlabeled exendin-4 resulted in self-inhibition (saturation) that only became statistically significant (P<0.05) when the results of four experiments were combined; this suggests a possible limit to the amount of peripherally administered exendin-4 that can reach the brain after injection of high doses. CONCLUSION: The results indicate that exendin-4 is well conformed for exerting central effects involved in the control of obesity.

Asunto(s)

Encéfalo/metabolismo , Péptidos/farmacocinética , Ponzoñas , Animales , Barrera Hematoencefálica , Cromatografía Líquida de Alta Presión/métodos , Evaluación Preclínica de Medicamentos , Exenatida , Enlace de Hidrógeno , Inyecciones Intravenosas , Masculino , Ratones , Ratones Endogámicos , Octanoles , Péptidos/sangre , Péptidos/química

RESUMEN

Endomorphin-2 is a newly discovered endogenous opioid peptide with high affinity and selectivity for the micro-opioid receptor, and potent analgesic activity, particularly in the spinal cord. Using immunoelectron microscopy, we examined the ultrastructure of the endomorphin-2-like immunoreactive processes and their synaptic relationships in the spinal cord. Endomorphin-2-like immunopositive dense-cored vesicles were observed in many axon terminals, and, in a few cases, were observed together with immunonegative dense-cored vesicles. Immunopositive axons with or without myelination were also observed. The endomorphin-2-like immunoreactive axon terminals formed synapses with both immunopositive and immunonegative processes. Most synapses were asymmetrical, but symmetrical synapses were also found. Examples of axo-dendritic, axo-somatic and axo-axonic contacts were observed. This first demonstration of the ultrastructure and synaptic relationships of endomorphin-2-like immunoreactive axon terminals in the spinal cord dorsal horn provides morphological evidence that this peptide functions as a transmitter regulating pain processes.

Asunto(s)

Microscopía Inmunoelectrónica , Oligopéptidos/análisis , Células del Asta Posterior/química , Células del Asta Posterior/ultraestructura , Animales , Vértebras Cervicales , Masculino , Oligopéptidos/inmunología , Terminales Presinápticos/química , Terminales Presinápticos/ultraestructura , Ratas , Ratas Wistar , Receptores Opioides mu/agonistas , Sinapsis/química , Sinapsis/ultraestructura

RESUMEN

Galanin-like peptide (GALP) was recently isolated from the hypothalamus, where its expression is influenced by leptin and food deprivation. Since leptin crosses the blood-brain barrier (BBB) by a saturable transport system that is downregulated by fasting, we examined the effect of leptin and fasting on the entry of GALP into mouse brain. Multiple-time regression analysis showed that the basal influx of 125I-GALP from blood was rapid (K(i) = 9.49 +/- 0.72 x 10(-4) ml/g x min). This influx was not affected by leptin but was significantly decreased by food deprivation for 24 or 48 h, accompanied by decreased immunoreactive plasma GALP at 48 h, but not at 24 h. By contrast, pretreatment of mice fasted for 24 h with glucose resulted in a significant increase in the blood-to-brain influx of GALP that was not accompanied by increased immunoreactive plasma GALP. HPLC showed that most of the GALP crossed the BBB in an intact form, and capillary depletion studies showed that more than 93% of the GALP crossing entered the parenchyma of the brain rather than being bound to the endothelial cells of the capillaries composing the BBB or being reversibly associated with the vasculature. Efflux of 125I-GALP occurred at the rate of the normal reabsorption of CSF, and the octanol-buffer partition coefficient showed insufficient lipophilicity to explain the fast rate of influx. When 125I-GALP was perfused in blood-free buffer, the self-inhibition characteristic of a saturable transport system was evident even though capillary gel electrophoresis showed GALP aggregating as a trimer. Capillary zone electrophoresis showed protein binding of GALP in serum, perhaps facilitating its interactions at the BBB. In particular, these studies show for the first time (1) that immunoreactive GALP is present in blood where (2) its concentrations are reduced by food deprivation, and (3) that there is a rapid blood-to-brain influx of intact GALP (4) which is decreased by fasting and (5) increased by pretreatment with glucose.

Asunto(s)

Encéfalo/metabolismo , Privación de Alimentos/fisiología , Proteínas del Tejido Nervioso/farmacocinética , Animales , Barrera Hematoencefálica , Tampones (Química) , Capilares/metabolismo , Cromatografía , Cromatografía Líquida de Alta Presión , Relación Dosis-Respuesta a Droga , Electroforesis , Péptido Similar a Galanina , Glucosa/farmacología , Inyecciones Intravenosas , Masculino , Ratones , Ratones Endogámicos ICR , Proteínas del Tejido Nervioso/administración & dosificación , Proteínas del Tejido Nervioso/sangre , Perfusión , Radioinmunoensayo

RESUMEN

Leptin regulates body adiposity by decreasing feeding and increasing thermogenesis. Obese humans and some obese rodents are resistant to peripherally administered leptin, suggesting a defect in the transport of leptin across the blood-brain barrier (BBB). Defective transport of exogenous leptin occurs in some models of obesity, but in other models transport is normal. This shows that factors other than obesity are associated with impairment of leptin transport across the BBB. In order to further investigate these factors, we determined leptin transport in rats made obese by lesioning of the ventromedial hypothalamus (VMH), paraventricular nucleus (PVN), or posterodorsal amygdala (PDA). These regions all contain leptin receptors and lesions there induce obesity and hyperleptinemia and alter the levels of many feeding hormones which might participate in leptin transporter regulation. We measured the uptake of radioactively labeled leptin by the BBB by multiple-time regression analysis which divides uptake into a reversible phase (Vi, e.g., receptor/transporter binding to the brain endothelial cell) and an irreversible phase (Ki, complete transport across the BBB). Leptin uptake was not affected in rats with VMH lesions. No significant change occurred in the entry rate (Ki) for any group, although Ki declined by over 35% in rats with PVN lesions. Decreased uptake was observed in rats with PVN lesions and with PDA lesions. This was primarily due to a reduced Vi (about 21% for the PDA). This decreased uptake is most likely explained by decreased binding of leptin to the brain endothelial cell, which could be because of decreased binding by either receptors or transporters. This suggests that some of the feeding hormones controlled by the PVN and PDA may participate in regulating leptin uptake by the BBB.

Asunto(s)

Amígdala del Cerebelo/metabolismo , Barrera Hematoencefálica/fisiología , Hipotálamo/metabolismo , Leptina/metabolismo , Obesidad/metabolismo , Núcleo Hipotalámico Paraventricular/metabolismo , Amígdala del Cerebelo/lesiones , Animales , Femenino , Hipotálamo/lesiones , Obesidad/etiología , Núcleo Hipotalámico Paraventricular/lesiones , Ratas , Ratas Long-Evans , Aumento de Peso

RESUMEN

Neuropeptides 2001, 2nd Joint Meeting of the European Neuropeptide Club and the American Summer Neuropeptide Conference (11th Annual Meeting). 6-11 May 2001 with Satellite Symposium, Israeli-French Symposium, Israel Ministry of Science, Culture and Sport, 6 May 2001, held at Maale Hachmicha and Tel Aviv University, Israel.

Asunto(s)

Química Encefálica/fisiología , Neuropéptidos/fisiología , Animales

RESUMEN

This study was designed to determine the safety, efficacy and pharmacokinetics of the antidepressant netamiftide (previously designated name: INN 00835) after 5 or 10 daily doses administered to patients diagnosed with major depression. Netamiftide was administered subcutaneously at a fixed dose of 18 mg/patient per day. Of the 55 enrolled patients, 22 were dosed for 10 days with drug, 11 for 5 days with drug followed by 5 days with placebo and 22 for 10 days with placebo only. The effect of treatment with netamiftide was evaluated by the following psychometric tests: Hamilton Depression Rating, Montgomery-Asberg Depression Rating Scale, Carroll Self-Rating Depression and Clinical Global Impression scales. None of the patients experienced significant adverse effects. A pharmacodynamic correlation (P < 0.05) was found between plasma drug concentrations and response to treatment. Highest plasma concentrations (Cmax) of netamiftide averaging 45.7 ng/ml were observed at 0.25 h after dosing. There were 89% responders in the group with Cmax > or = 45.7 ng/ml (minimum therapeutic concentration) versus 40% in the group with Cmax < 45.7 ng/ml. Onset of action was observed within 48 h after treatment, peak effect was observed at approximately 1 week after treatment and efficacy lasted during a 4-week follow-up period. Netamiftide is a promising antidepressant with rapid onset of action and with an excellent safety profile.

Asunto(s)

Antidepresivos/uso terapéutico , Trastorno Depresivo/tratamiento farmacológico , Oligopéptidos/uso terapéutico , Adolescente , Adulto , Anciano , Antidepresivos/efectos adversos , Antidepresivos/sangre , Trastorno Depresivo/psicología , Método Doble Ciego , Femenino , Humanos , Masculino , Persona de Mediana Edad , Oligopéptidos/efectos adversos , Oligopéptidos/sangre , Pacientes Ambulatorios , Proyectos Piloto , Escalas de Valoración Psiquiátrica

RESUMEN

Two highly selective mu-opioid receptor agonists, endomorphin-1 and endomorphin-2, have been identified and postulated to be endogenous ligands for mu-opioid receptors. Intrathecal (i.t.) administration of endomorphin-1 and endomorphin-2 at doses from 0.039 to 5 nmol dose-dependently produced antinociception with the paw-withdrawal test. The paw-withdrawal inhibition rapidly reached its peak at 1 min, rapidly declined and returned to the pre-injection levels in 20 min. The inhibition of the paw-withdrawal responses to endomorphin-1 and endomorphin-2 at a dose of 5 nmol observed at 1 and 5 min after injection was blocked by pretreatment with a non-selective opioid receptor antagonist naloxone (1 mg/kg, s.c.). The antinociceptive effect of endomorphin-2 was more sensitive to the mu (1)-opioid receptor antagonist, naloxonazine than that of endomorphin-1. The endomorphin-2-induced paw-withdrawal inhibition at both 1 and 5 min after injection was blocked by pretreatment with kappa-opioid receptor antagonist nor-binaltorphimine (10 mg/kg, s.c.) or the delta(2)-opioid receptor antagonist naltriben (0.6 mg/kg, s.c.) but not the delta(1)-opioid receptor antagonist 7-benzylidine naltrexone (BNTX) (0.6 mg/kg s.c.). In contrast, the paw-withdrawal inhibition induced by endomorphin-1 observed at both 1 and 5 min after injection was not blocked by naloxonazine (35 mg/kg, s.c.), nor-binaltorphimine (10 mg/kg, s.c.), naltriben (0.6 mg/kg, s.c.) or BNTX (0.6 mg/kg s.c.). The endomorphin-2-induced paw-withdrawal inhibition was blocked by the pretreatment with an antiserum against dynorphin A-(1-17) or [Met(5)]enkephalin, but not by antiserum against dynorphin B-(1-13). Pretreatment with these antisera did not affect the endomorphin-1-induced paw-withdrawal inhibition. Our results indicate that endomorphin-2 given i.t. produces its antinociceptive effects via the stimulation of mu (1)-opioid receptors (naloxonazine-sensitive site) in the spinal cord. The antinociception induced by endomophin-2 contains additional components, which are mediated by the release of dynorphin A-(1-17) and [Met(5)]enkephalin which subsequently act on kappa-opioid receptors and delta(2)-opioid receptors to produce antinociception.

Asunto(s)

Analgésicos/farmacología , Naloxona/análogos & derivados , Naltrexona/análogos & derivados , Oligopéptidos/farmacología , Animales , Compuestos de Bencilideno/farmacología , Relación Dosis-Respuesta a Droga , Dinorfinas/inmunología , Encefalina Ala(2)-MeFe(4)-Gli(5)/farmacología , Encefalina Leucina/inmunología , Encefalina Metionina/inmunología , Sueros Inmunes/farmacología , Inyecciones Espinales , Inyecciones Subcutáneas , Masculino , Ratones , Naloxona/farmacología , Naltrexona/farmacología , Antagonistas de Narcóticos/farmacología , Dolor/prevención & control , Dimensión del Dolor , Umbral del Dolor/efectos de los fármacos , Fragmentos de Péptidos/inmunología , Factores de Tiempo

RESUMEN

Loss of ovarian function, such as occurs with menopause in human beings and ovariectomy in rodents, results in weight gain. Using multiple-time regression analysis, a sensitive technique for quantifying blood-to-brain transport of peptides and polypeptides, we found that mice ovariectomized for at least 5 weeks had markedly reduced entry of the satiety factor leptin into brain. The rate of entry of leptin into brain remained reduced half a year later. The results suggest that the weight gain resulting from loss of ovarian function could be explained by decreased transport of leptin into the brain.

Asunto(s)

Química Encefálica/fisiología , Leptina/metabolismo , Ovario/fisiología , Animales , Barrera Hematoencefálica , Femenino , Radioisótopos de Yodo , Ratones , Ovariectomía , Aumento de Peso/fisiología

RESUMEN

Opiate-modulating tetrapeptides such as tyrosine-melanocyte-stimulating hormone-release inhibiting factor-1 (Tyr-MIF-1; Tyr-Pro-Leu-Gly-NH2) and Tyr-W-MIF-1 (Tyr-Pro-Trp-Gly-NH2) are saturably transported from brain to blood. We examined whether two recently described endogenous opiate tetrapeptides with similar structures, the mu-specific endomorphins, also are transported across the blood-brain barrier (BBB). We found that the efflux rates of endomorphin-1 (Tyr-Pro-Trp-Phe-NH2) and endomorphin-2 (Tyr-Pro-Phe-Phe-NH2) were each self-inhibited by an excess of the respective endomorphin, thereby defining saturable transport. Cross-inhibition of the transport of each endomorphin by the other indicated shared transport. By contrast, no inhibition of the efflux of either endomorphin resulted from coadministration of Tyr-MIF-1, indicating that peptide transport system-1 (PTS-1) was not involved. Tyr-W-MIF-1, which is partially transported by PTS-1, significantly (P<0.01) decreased the transport of endomorphin-1 and tended (P=0.051) to decrease the transport of endomorphin-2, consistent with its role as both an opiate and antiopiate. Although involved in modulation of pain, coinjection of calcitonin gene-related peptide or constriction of the sciatic nerve did not appear to inhibit endomorphin efflux. Thus, the results demonstrate the existence of a new efflux system across the BBB which saturably transports endomorphins from brain to blood.

Asunto(s)

Barrera Hematoencefálica/fisiología , Encéfalo/metabolismo , Proteínas Portadoras/metabolismo , Hormona Inhibidora de la Liberación de MSH/análogos & derivados , Oligopéptidos/metabolismo , Receptores Opioides mu/metabolismo , Animales , Unión Competitiva/efectos de los fármacos , Unión Competitiva/fisiología , Barrera Hematoencefálica/efectos de los fármacos , Encéfalo/efectos de los fármacos , Péptido Relacionado con Gen de Calcitonina/metabolismo , Péptido Relacionado con Gen de Calcitonina/farmacología , Proteínas Portadoras/efectos de los fármacos , Radioisótopos de Yodo/farmacocinética , Ligadura , Hormona Inhibidora de la Liberación de MSH/metabolismo , Hormona Inhibidora de la Liberación de MSH/farmacocinética , Masculino , Proteínas de Transporte de Membrana/efectos de los fármacos , Proteínas de Transporte de Membrana/metabolismo , Ratones , Ratones Endogámicos ICR , Oligopéptidos/farmacocinética , Dolor/metabolismo , Dolor/fisiopatología , Ensayo de Unión Radioligante , Receptores Opioides mu/efectos de los fármacos , Nervio Ciático/lesiones , Nervio Ciático/fisiopatología , Nervio Ciático/cirugía

RESUMEN

The blood-brain barrier (BBB), composed of the microvessels of cerebral capillary endothelial cells, regulates the passage of peptides into the brain in several ways, mainly by saturable transport or passive diffusion. Here we describe an additional mechanism by which this regulatory function can occur. Cerebral microvessels were isolated from different regions of the brain and incubated with the mu-opiate selective endomorphin-1 (Tyr-Pro-Trp-Phe-NH2) or the opiate-modulating Tyr-MIF-1 (Tyr-Pro-Leu-Gly-NH2), both tetrapeptides selectively tritiated at the Pro. Degradation was determined by HPLC. For both peptides, the metabolism by microvessels from the cerebral cortex was much greater than that by microvessels from the hypothalamus or pons. For endomorphin-1, the least degradation was in the pons; for Tyr-MIF-1 there was no difference in metabolism by microvessels from the pons or hypothalamus. The results show a novel mechanism at the BBB by which the BBB can selectively regulate the activity of different peptides in different regions of the brain.

Asunto(s)

Barrera Hematoencefálica/fisiología , Corteza Cerebral/metabolismo , Hormona Inhibidora de la Liberación de MSH/metabolismo , Oligopéptidos/metabolismo , Animales , Capilares/metabolismo , Corteza Cerebral/irrigación sanguínea , Cromatografía Líquida de Alta Presión , Endotelio Vascular/metabolismo , Hipotálamo/irrigación sanguínea , Hipotálamo/metabolismo , Hormona Inhibidora de la Liberación de MSH/análogos & derivados , Masculino , Puente/irrigación sanguínea , Puente/metabolismo , Ratas , Ratas Sprague-Dawley

RESUMEN

Bradykinin is one of the key molecules involved in the disruption of the blood-brain barrier and blood-spinal cord barrier occurring after spinal cord injury (SCI). Previously we have shown a biphasic opening of the blood-spinal cord barrier as well as increased transport of tumor necrosis factor-alpha (TNFalpha) after SCI by compression of the lumbar spinal cord in mice. To evaluate the role of bradykinin in the two phases of blood-spinal cord barrier disruption, we pretreated mice with a potent bradykinin antagonist, the decapeptide B9430, before SCI. Our results show that B9430 decreased the general blood-spinal cord barrier disruption occurring immediately after SCI but failed to affect the delayed opening of the blood-spinal cord barrier observed 72 h after SCI. By contrast, the entry of TNFalpha after SCI was not affected by B9430 treatment. We conclude that bradykinin is involved in the early phase of blood-spinal cord barrier disruption, with B9430 non-selectively blocking this early disruption without affecting the selective transport system for TNFalpha. This indicates the therapeutic potential of bradykinin antagonists in ameliorating tissue damage induced by SCI.

Asunto(s)

Barrera Hematoencefálica/efectos de los fármacos , Bradiquinina/antagonistas & inhibidores , Bradiquinina/farmacología , Traumatismos de la Médula Espinal/tratamiento farmacológico , Médula Espinal/efectos de los fármacos , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Barrera Hematoencefálica/fisiología , Bradiquinina/análogos & derivados , Bradiquinina/metabolismo , Permeabilidad de la Membrana Celular/efectos de los fármacos , Permeabilidad de la Membrana Celular/fisiología , Radioisótopos de Yodo/farmacocinética , Masculino , Ratones , Ratones Endogámicos ICR , Médula Espinal/metabolismo , Médula Espinal/fisiopatología , Traumatismos de la Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/fisiopatología , Agregado de Albúmina Marcado con Tecnecio Tc 99m/farmacocinética , Factores de Tiempo

RESUMEN

Nerve injury often leads to chronic, sometimes excruciating, pain. The mechanisms contributing to this syndrome include neurochemical plasticity in neurons involved in the earliest stages of pain transmission. Endomorphin-2 (Tyr-Pro-Phe-Phe-NH(2)) is an endogenous morphine-like substance that binds to the mu-opioid receptor with high affinity and selectivity. Endomorphin-2-like immunoreactivity (LI) is present in the superficial layers of the dorsal horn in the spinal cord and in primary afferents, suggesting a role for this peptide in pain transmission. To determine whether spinal endomorphin-2-LI is altered in an animal model of chronic pain, the left sciatic nerve of Swiss Webster and ICR mice was ligated in a modified Seltzer model of nerve injury. Changes in endomorphin-2-LI were assessed by immunocytochemistry at 2, 4 and 14 days after nerve injury. The side of the spinal cord ipsilateral to the nerve injury exhibited a dramatic decrease in endomorphin-2-LI relative to the contralateral side and to control animals. The change was restricted to the medial dorsal horn in the lumbar segments innervated by the sciatic nerve. Substance P-LI showed a small decrease, while calcitonin gene-related peptide-LI was unchanged. Both thermal hyperalgesia, as evidenced by significantly decreased paw withdrawal latencies, and decreased endomorphin-2-LI were observed within 2 days of injury and were most pronounced at 2 weeks after injury. The decrease in endomorphin-2-LI during the development of chronic pain is consistent with the loss of an inhibitory influence on pain transmission. These results provide the first evidence that reduction of an endogenous opioid in primary afferents is associated with injury-induced chronic pain.

Asunto(s)

Regulación hacia Abajo/fisiología , Neuralgia/metabolismo , Oligopéptidos/metabolismo , Enfermedades del Sistema Nervioso Periférico/metabolismo , Nervio Ciático/lesiones , Nervio Ciático/metabolismo , Animales , Péptido Relacionado con Gen de Calcitonina/metabolismo , Enfermedad Crónica , Lateralidad Funcional/fisiología , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatología , Inmunohistoquímica , Ligadura , Masculino , Ratones , Ratones Endogámicos ICR , Compresión Nerviosa , Neuralgia/fisiopatología , Dimensión del Dolor , Umbral del Dolor/fisiología , Enfermedades del Sistema Nervioso Periférico/fisiopatología , Células del Asta Posterior/citología , Células del Asta Posterior/metabolismo , Tiempo de Reacción/fisiología , Nervio Ciático/cirugía , Sustancia P/metabolismo

RESUMEN

The dynamic changes of the blood-brain barrier and blood-spinal cord barrier (BBB) are an important part of the CNS response to injury. This study addresses the permeability of the BBB in the acute phase of spinal cord injury (SCI) to the thoracic region. SCI by compression or by complete transection was generated in mice. BBB disruption was evaluated by spinal cord uptake of radiolabeled albumin. The BBB of the thoracic spinal cord was disrupted immediately after compression injury, lasting for 2 days. This was followed by a delayed permeability increase in the cervical spinal cord beginning 3 days after injury. After transection, BBB disruption was limited to the thoracic spinal cord and was present only immediately postinjury. The entry of TNFalpha not only was increased at the time of BBB disruption, following the same pattern, but also had secondary changes after the BBB permeability to albumin had returned to normal. The increase of TNFalpha entry, best explained by upregulation of the specific transport system for TNFalpha, was pronounced in the lumbar spinal cord as well as the thoracic region, and followed a different time course after the two types of injury. Integrating our results with those of the literature regarding the roles of inflammatory responses and the effects of TNFalpha in spinal cord regeneration, we conclude that the time-, region-, and lesion-specificity of the upregulation of TNFalpha transport is part of the regulatory changes at the BBB in response to SCI.

Asunto(s)

Barrera Hematoencefálica , Traumatismos de la Médula Espinal/fisiopatología , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Modelos Animales de Enfermedad , Cinética , Ratones , Ratones Endogámicos ICR , Compuestos de Organotecnecio/farmacocinética , Radiofármacos/sangre , Radiofármacos/farmacocinética , Albúmina Sérica/farmacocinética , Traumatismos de la Médula Espinal/patología , Tórax , Factores de Tiempo

RESUMEN

The blood-brain barrier (BBB) regulates the amount of peripherally produced leptin reaching the brain. Knowing that the blood concentration of leptin has a circadian rhythm, we investigated whether the influx of leptin at the BBB followed the same pattern in three main sets of experiments. (a): The entry of 125I-leptin from blood to brain was measured in mice every 4 h, as indicated by the influx rate of 125I-leptin 1-10 min after an iv bolus injection. The blood concentration of endogenous leptin was measured at the same times. Blood leptin concentrations were higher at night and early morning (peak at 0800 h) and lower during the day (nadir at 1600 h). By contrast, the influx of 125I-leptin was fastest at 2000 h and slowest at 0400 h. Addition of unlabeled leptin (1 microg/mouse) significantly decreased the influx rate of 125I-leptin at all time points, indicating saturability of the transport system. The unlabeled leptin also abolished the diurnal variation of the influx of 125I-leptin. (b): The entry of 125I-leptin into spinal cord was faster than that into brain and showed a different diurnal pattern. The greatest influx occurred at 2400 h and the slowest at 0800 h. In spinal cord, unlike brain, unlabeled leptin (1 microg/mouse) neither inhibited the influx of 125I-leptin nor abolished the diurnal rhythm. (c): Higher concentrations of unlabeled leptin (5 microg/mouse) inhibited the uptake of 125I-leptin in spinal cord as well as in brain, but not in muscle. This experiment measured uptake 10 min after iv injection at 0600 h (beginning of the light cycle) and 1800 h (beginning of the dark cycle). Thus, influx of 125I-leptin into the CNS shows diurnal variation, indicating a circadian rhythm in the transport system at the BBB, saturation of the leptin transport system shows differences between the brain and spinal cord, and blood concentrations of leptin suggest that partial saturation of the transport system occurs at physiological concentrations of circulating leptin, contributing to the differing diurnal patterns in brain and spinal cord. Together, the results show that the BBB is actively involved in the neuroendocrine regulation of feeding behavior.

Asunto(s)

Barrera Hematoencefálica , Encéfalo/metabolismo , Ritmo Circadiano , Leptina/metabolismo , Animales , Transporte Biológico , Luz , Masculino , Ratones , Ratones Endogámicos ICR

RESUMEN

A saturable blood-to-brain transport system for leptin across the blood-brain barrier (BBB) has been observed in vivo. Since the main component of the non-fenestrated microvessels of the BBB is the endothelial cell, we established an in vitro culture system of these cerebrovascular cells to study leptin transport and to determine whether the self-inhibition of leptin transport characteristic of a saturable system occurs at this level. The results show that 125I-leptin crossed from the luminal to abluminal side of a monolayer of cerebral microvessel cells significantly faster than the albumin and lactalbumin controls. This transport of 125I-leptin across an in vitro BBB was significantly faster than in the opposite direction and was dose-relatedly inhibited by the addition of unlabeled leptin. Thus, the results establish that the saturable transport system for leptin across the BBB occurs at the level of the endothelial cells of the BBB.

Asunto(s)

Barrera Hematoencefálica/fisiología , Circulación Cerebrovascular/fisiología , Endotelio Vascular/fisiología , Leptina/metabolismo , Microcirculación/fisiología , Animales , Transporte Biológico , Membrana Celular/metabolismo , Células Cultivadas , Cinética , Lactalbúmina/metabolismo , Leptina/sangre , Masculino , Ratas , Ratas Sprague-Dawley , Albúmina Sérica/metabolismo

RESUMEN

Although urocortin is a potent inhibitor of food ingestion after peripheral administration, it was recently shown that under normal conditions this peptide crosses the blood-brain barrier (BBB) at a very slow rate. We examined whether hyperglycemia could stimulate the rate of entry (K(i)) of (125)I-urocortin into the mouse brain. In euglycemic mice, (125)I-urocortin injected iv entered the brain at a rate similar to that of the vascular marker (99m)Tc-albumin. However, injection of glucose (3 g/kg, ip) 0.5, 1, or 2 h before the (125)I-urocortin greatly increased the influx of urocortin. Without the glucose, the self-inhibition characteristic of a saturable transport system was not apparent. Self-inhibition could be demonstrated after the glucose injection, indicating activation of a transport system for urocortin that was saturable. Injection of insulin (10 U/kg, ip) 1 or 2 h before the (125)I-urocortin decreased the K(i). Thus, the entry of urocortin into brain can be activated by changes in the concentration of blood glucose, illustrating the responsiveness of the BBB to regulatory influences.

Asunto(s)

Encéfalo/metabolismo , Hormona Liberadora de Corticotropina/efectos de los fármacos , Hormona Liberadora de Corticotropina/farmacocinética , Glucosa/agonistas , Insulina/farmacología , Animales , Glucemia/metabolismo , Barrera Hematoencefálica/efectos de los fármacos , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Hiperglucemia/metabolismo , Masculino , Ratones , Ratones Endogámicos ICR , Estreptozocina , Urocortinas

RESUMEN

Since fasting is one of the few factors found to change the rate of entry of leptin into brain, we used multiple-time regression analysis to study the effects of pretreatment with glucose or insulin on leptin transport across the blood-brain barrier (BBB). Two hours after intraperitoneal injection of glucose (3 g/kg), there was a statistically significant increase in the entry rate (K(i)) of leptin in fasted (from 4.91 +/- 0.70 x 10(-4) ml/g x min to 9.03 +/- 1.00 x 10(-4) ml/g x min) but not (p = 0.15) in nonfasted normal (from 4.90 +/- 1.21 x 10(-4) ml/g x min to 6.42 +/- 1.79 x 10(-4) ml/g x min) or fasted streptozotocin (STZ)-treated diabetic mice (from 4.043 +/- 0.959 x 10(-4) ml/g min to 5.395 +/- 1.355 x 10(-4) ml/g min). Insulin (10 U/kg) increased leptin influx in fasted (from 4.77 +/- 0.26 x 10(-4) ml/g x min to 10.6 +/- 0.15 x 10(-4) ml/g x min at 0.5 h) and nonfasted (from 4.64 +/- 0.75 x 10(-4) ml/g x min to 7.46 +/- 1.48 x 10(-4) ml/g x min at 0.5 h) normal mice, but not in STZ-diabetic mice deficient in insulin (and leptin), even though basal concentrations of glucose were similarly increased in the nonfasted normal and STZ-treated mice. Moreover, the basal rate of leptin influx was the same in overnight fasted normal mice, nonfasted normal mice and STZ-diabetic mice. The results indicate that glucose and insulin can increase leptin transport, but they probably are not the principal factors responsible for the regulatory effect of the BBB on leptin entry into the brain.

Asunto(s)

Barrera Hematoencefálica/efectos de los fármacos , Diabetes Mellitus Experimental/metabolismo , Glucosa/farmacología , Hipoglucemiantes/farmacología , Insulina/farmacología , Leptina/metabolismo , Animales , Glucemia/metabolismo , Glucosa/administración & dosificación , Hipoglucemiantes/administración & dosificación , Inyecciones Intraperitoneales , Inyecciones Intravenosas , Insulina/administración & dosificación , Insulina/sangre , Leptina/farmacocinética , Ratones , Ratones Endogámicos ICR , Agregado de Albúmina Marcado con Tecnecio Tc 99m