RESUMO
Disturbances in respiration are common and debilitating features of Rett syndrome (RTT). A previous study showed that the 5-HT1a receptor agonist (R)-(+)-8-hydroxy-dipropyl-2-aminotetralin hydrobromide (8-OH-DPAT) significantly reduced the incidence of apnea and the irregular breathing pattern in a mouse model of the disorder. 8-OH-DPAT, however, is not available for clinical practice. Sarizotan, a full 5-HT1a agonist and a dopamine D2-like agonist/partial agonist, has been used in clinical trials for the treatment of l-dopa-induced dyskinesia. The purpose of this study was to evaluate the effects of sarizotan on respiration and locomotion in mouse models of RTT. Studies were performed in Bird and Jaenisch strains of methyl-CpG-binding protein 2--deficient heterozygous female and Jaenisch strain Mecp2 null male mice and in knock-in heterozygous female mice of a common nonsense mutation (R168X). Respiratory pattern was determined with body plethysmography, and locomotion was determined with open-field recording. Sarizotan or vehicle was administered 20 minutes before a 30-minute recording of respiratory pattern or motor behavior. In separate studies, a crossover design was used to administer the drug for 7 and for 14 days. Sarizotan reduced the incidence of apnea in all three RTT mouse models to approximately 15% of their pretreatment levels. The irregular breathing pattern was corrected to that of wild-type littermates. When administered for 7 or 14 days, apnea decreased to 25 to 33% of the incidence seen with vehicle. This study indicates that the clinically approved drug sarizotan is an effective treatment for respiratory disorders in mouse models of RTT.
Assuntos
Respiração/efeitos dos fármacos , Síndrome de Rett/tratamento farmacológico , Agonistas do Receptor 5-HT1 de Serotonina/farmacologia , Animais , Modelos Animais de Doenças , Feminino , Locomoção/efeitos dos fármacos , Locomoção/genética , Masculino , Proteína 2 de Ligação a Metil-CpG/genética , Proteína 2 de Ligação a Metil-CpG/metabolismo , Camundongos , Camundongos Knockout , Compostos Orgânicos/farmacologia , Receptor 5-HT1A de Serotonina/genética , Receptor 5-HT1A de Serotonina/metabolismo , Receptores de Dopamina D2/agonistas , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo , Respiração/genética , Síndrome de Rett/genética , Síndrome de Rett/metabolismoRESUMO
Mice deficient in the transcription factor methyl-CpG-binding protein 2 (Mecp2), a mouse model of Rett syndrome, display reduced CO2 chemosensitivity, which may contribute to their breathing abnormalities. In addition, patients with Rett syndrome and male mice that are null for Mecp2 show reduced levels of brain serotonin (5-HT). Serotonin is known to play a role in central chemosensitivity, and we hypothesized that increasing the availability of 5-HT in this mouse model would improve their respiratory response to CO2. Here we determined the apnoeic threshold in heterozygous Mecp2-deficient female mice and examined the effects of blocking 5-HT reuptake on the CO2 response in Mecp2-null male mice. Studies were performed in B6.129P2(C)-Mecp2(τm1.1Bird) null males and heterozygous females. In an in situ preparation, seven of eight Mecp2-deficient heterozygous females showed arrest of phrenic nerve activity when arterial CO2 was lowered to 3%, whereas the wild-types maintained phrenic nerve amplitude at 53 ± 3% of maximal. In vivo plethysmography studies were used to determine CO2 chemosensitivity in null males. These mice were exposed sequentially to 1, 3 and 5% CO2. The percentage increase in minute ventilation in response to increased inspired CO2 was less in Mecp2(-/y) than in Mecp2(+/y) mice. Pretreatment with citalopram, a selective 5-HT reuptake inhibitor (2.5 mg kg(-1) i.p.), 40 min prior to CO2 exposure, in Mecp2(-/y) mice resulted in an improvement in CO2 chemosensitivity to wild-type levels. These results suggest that decreased 5-HT in Mecp2-deficient mice reduces CO2 chemosensitivity, and restoring 5-HT levels can reverse this effect.
Assuntos
Dióxido de Carbono/toxicidade , Proteína 2 de Ligação a Metil-CpG/deficiência , Transtornos Respiratórios/induzido quimicamente , Serotonina/administração & dosagem , Animais , Apneia/etiologia , Apneia/fisiopatologia , Encéfalo/metabolismo , Citalopram/uso terapêutico , Modelos Animais de Doenças , Feminino , Técnicas In Vitro , Masculino , Camundongos , Camundongos Knockout , Transtornos Respiratórios/tratamento farmacológico , Transtornos Respiratórios/fisiopatologia , Serotonina/deficiênciaRESUMO
Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X-linked gene methyl-CpG-binding protein 2 (Mecp2) that encodes a DNA binding protein involved in gene silencing. Periodic breathing (Cheyne-Stokes respiration) is commonly seen in RTT. Freely moving mice were studied with continuous recording of pleural pressure by telemetry. Episodes of periodic breathing in heterozygous Mecp2 deficient (Mecp2(+/-)) female mice (9.4 +/- 2.2 h(-1)) exceeded those in wild-type (Mecp2(+/+)) animals (2.5 +/- 0.4 h(-1)) (P = 0.010). Exposing Mecp2(+/-) animals to 40% oxygen increased the amount of periodic breathing from 118 +/- 25 s/30 min in air to 242 +/- 57 s/30 min (P = 0.001), and 12% oxygen tended to decrease it (67 +/- 29 s/30 min, P = 0.14). Relative hyperoxia and hypoxia did not affect the incidence of periodic breathing in Mecp2(+/+) animals. The ventilation/apnea ratio (V/A) was less at all levels of oxygen in heterozygous Mecp2(+/-) females compare with wild type (P = 0.003 to P < 0.001), indicating that their loop gain is larger. V/A in Mecp2(+/-) fell from 2.42 +/- 0.18 in normoxia to 1.82 +/- 0.17 in hyperoxia (P = 0.05) indicating an increase in loop gain with increased oxygen. Hyperoxia did not affect V/A in Mecp2(+/+) mice (3.73 +/- 0.28 vs. 3.5 +/- 0.28). These results show that periodic breathing in this mouse model of RTT is not dependent on enhanced peripheral chemoreceptor oxygen sensitivity. Rather, the breathing instability is of central origin.
Assuntos
Respiração de Cheyne-Stokes/fisiopatologia , Hiperóxia/fisiopatologia , Hipóxia/fisiopatologia , Proteína 2 de Ligação a Metil-CpG/metabolismo , Oxigênio/administração & dosagem , Ventilação Pulmonar/efeitos dos fármacos , Mecânica Respiratória/efeitos dos fármacos , Síndrome de Rett/complicações , Administração por Inalação , Animais , Dióxido de Carbono/administração & dosagem , Respiração de Cheyne-Stokes/etiologia , Respiração de Cheyne-Stokes/metabolismo , Modelos Animais de Doenças , Feminino , Hiperóxia/metabolismo , Hipóxia/metabolismo , Proteína 2 de Ligação a Metil-CpG/deficiência , Proteína 2 de Ligação a Metil-CpG/genética , Camundongos , Camundongos Transgênicos , Centro Respiratório/fisiopatologia , Síndrome de Rett/metabolismo , Síndrome de Rett/fisiopatologia , Telemetria , Fatores de TempoRESUMO
Methyl-CpG-binding protein 2 is a transcription factor that is involved in gene silencing. It is mutated in the majority of cases of Rett syndrome. This X-linked neurodevelopmental disorder is reported to involve abnormalities in autonomic cardiovascular regulation. As an initial step in understanding the basis for these abnormalities we have characterized autonomic cardiovascular function in Mecp2 deficient mice. Arterial pressure waves were recorded in freely moving animals using telemetry. Baseline blood pressure and pulse interval (PI) as well as indices of heart rate variability (HRV): standard deviation of PI (SDNN), range encompassing 90% of PIs (PI90) and standard deviation of adjacent PIs (SDSD) were similar in Mecp2(+/+) and Mecp2(+/-) animals. Spectral analysis of mean arterial pressure (MAP) and PI in the frequency domain showed similar relative power in low frequency 1 (LF1, 08-0.4 Hz), low frequency 2 (LF2, 0.4-1.0 Hz), middle frequency (MF, 1-3 Hz) and high frequency (HF, 3.0-10.0 Hz) bands. Autonomic blockade with atropine or propranolol as well as elevation in ambient temperature to 32 degrees C resulted in changes in blood pressure, PI and HRV that did not differ between the strains. Atropine, propranolol and elevated temperature resulted in similar changes in both MAP and PI spectral power. Baroreceptor function was tested using intravenous injections of nitroprusside followed by phenylephrine. Maximum gain was not different. These results do reveal any disturbance of autonomic cardiovascular regulation in the Mecp2 deficient mouse genotype.
Assuntos
Arritmias Cardíacas/genética , Doenças do Sistema Nervoso Autônomo/genética , Doenças Cardiovasculares/genética , Predisposição Genética para Doença/genética , Proteína 2 de Ligação a Metil-CpG/genética , Síndrome de Rett/genética , Antagonistas Adrenérgicos beta/farmacologia , Animais , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/fisiopatologia , Sistema Nervoso Autônomo/fisiopatologia , Doenças do Sistema Nervoso Autônomo/enzimologia , Doenças do Sistema Nervoso Autônomo/fisiopatologia , Barorreflexo/efeitos dos fármacos , Barorreflexo/fisiologia , Pressão Sanguínea/genética , Temperatura Corporal/fisiologia , Doenças Cardiovasculares/enzimologia , Doenças Cardiovasculares/fisiopatologia , Antagonistas Colinérgicos/farmacologia , Feminino , Coração/inervação , Coração/fisiopatologia , Sistema de Condução Cardíaco/fisiopatologia , Frequência Cardíaca/efeitos dos fármacos , Frequência Cardíaca/genética , Hipertermia Induzida , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Síndrome de Rett/complicações , Síndrome de Rett/enzimologiaRESUMO
Mice that are deficient in the transcription factor methyl-CpG-binding protein 2 (MeCP2) have a depressed hypercapnic ventilatory response (HCVR). The expression of MeCP2 can be selectively removed from astrocytes or neurons, thus offering a tool to dissect the role of this transcription factor in astrocytes from that in neurons. Studies were carried out in the progeny of mice that were a cross between those harboring a tamoxifen (TAM)-inducible Cre recombinase transgene driven by the human astrocytic glial fibrillary acidic protein (hGFAP) promoter, or Cre recombinase under control of the synapsin promoter, with mice containing a Cre-excisable exon III in the Mecp2 gene. The TAM-conditional excision of the Mecp2 exon allowed the respiratory CO2 response to be studied in the same animals before and after selective depletion of MeCP2 in astrocytes. Immunohistochemistry showed that following TAM treatment only â¼20% of GFAP-labeled cells in the retrotrapazoid nucleus and in the raphé magnus were positive for MeCP2. The slope of the relative increase in minute ventilation as a function of 1, 3, and 5% inspired CO2 was depressed in mice with depleted astrocyte MeCP2 compared with wild-type littermates. In contrast, selective depletion of MeCP2 in neurons did not significantly affect slope. While neurons which constitute the respiratory network ultimately determine the ventilatory response to CO2, this study demonstrates that loss of MeCP2 in astrocytes alone is sufficient to result in a dramatic attenuation of the HCVR. We propose that the glial contribution to HCVR is under the control of the MeCP2 gene.
Assuntos
Astrócitos/metabolismo , Hipercapnia/metabolismo , Proteína 2 de Ligação a Metil-CpG/metabolismo , Animais , Dióxido de Carbono/metabolismo , Células Cultivadas , Proteína Glial Fibrilar Ácida/metabolismo , Camundongos , Neuroglia/metabolismo , Neurônios/metabolismoRESUMO
BACKGROUND: Recent reports have shown that there are developmental changes in the ventilatory response to hypercapnia in the rat. These are characterized by an initial large response to carbon dioxide immediately after birth followed by a decline with a trough at one week of age, followed by a return in sensitivity. A second abnormality is seen at postnatal day 5 (P5) rats in that they cannot maintain the increase in frequency for 5 min of hypercapnia. In mice lacking GAD65 the release of GABA during sustained synaptic activation is reduced. We hypothesized that this developmental pattern would be present in the mouse which is also less mature at birth and that GABA mediates this relative respiratory depression. METHODS: In awake C57BL/6J and GAD65-/- mice the ventilatory response to 5% carbon dioxide (CO2) was examined at P2, P4, P6, P7, P12.5, P14.5 and P21.5, using body plethysmography. RESULTS: Minute ventilation (VE) relative to baseline during hypercapnia from P2 through P7 was generally less than from P12.5 onwards, but there was no trough as in the rat. Breaking VE down into its two components showed that tidal volume remained elevated for the 5 min of exposure to 5% CO2. At P6, but not at other ages, respiratory frequency declined with time and at 5 min was less that at 2 and 3 min. GAD65-/- animals at P6 showed a sustained increase in respiratory rate for the five mins exposure to CO2. CONCLUSION: These results show, that in contrast to the rat, mice do not show a decline in minute ventilatory response to CO2 at one week of age. Similar to the rat at P5, mice at P6 are unable to sustain an increase in CO2 induced respiratory frequency and GAD65 contributes to this fall off.
Assuntos
Envelhecimento , Glutamato Descarboxilase/deficiência , Glutamato Descarboxilase/metabolismo , Hipercapnia/fisiopatologia , Isoenzimas/deficiência , Isoenzimas/metabolismo , Mecânica Respiratória , Volume de Ventilação Pulmonar , Animais , Animais Recém-Nascidos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Isoformas de Proteínas/deficiência , Isoformas de Proteínas/metabolismo , Ventilação PulmonarRESUMO
Rett syndrome is a neurological disorder caused by loss of function mutations in the gene that encodes the DNA binding protein methyl-CpG-binding protein 2 (Mecp2). A prominent feature of the syndrome is disturbances in respiration characterized by frequent apnea and an irregular interbreath cycle. 8-Hydroxy-2-dipropylaminotetralin has been shown to positively modulate these disturbances (Abdala AP, Dutschmann M, Bissonnette JM, Paton JF, Proc Natl Acad Sci U S A 107: 18208-18213, 2010), but the mode of action is not understood. Here we show that the selective 5-HT1a biased agonist 3-chloro-4-fluorophenyl-(4-fluoro-4-{[(5-methylpyrimidin-2-ylmethyl)-amino]-methyl}-piperidin-1-yl)-methanone (F15599) decreases apnea and corrects irregularity in both heterozygous Mecp2-deficient female and in Mecp2 null male mice. In whole cell voltage-clamp recordings from dorsal raphe neurons, F15599 potently induced an outward current, which was blocked by barium, reversed at the potassium equilibrium potential, and was antagonized by the 5-HT1a antagonist WAY100135. This is consistent with somatodendritic 5-HT1a receptor-mediated activation of G protein-coupled inwardly rectifying potassium channels (GIRK). In contrast, F15599 did not activate 5-HT1b/d receptors that mediate inhibition of glutamate release from terminals in the nucleus accumbens by a presynaptic mechanism. Thus F15599 activated somatodendritic 5-HT1a autoreceptors, but not axonal 5-HT1b/d receptors. In unanesthetized Mecp2-deficient heterozygous female mice, F15599 reduced apnea in a dose-dependent manner with maximal effect of 74.5 ± 6.9% at 0.1 mg/kg and improved breath irrregularity. Similarly, in Mecp2 null male mice, apnea was reduced by 62 ± 6.6% at 0.25 mg/kg, and breathing became regular. The results indicate respiration is improved with a 5-HT1a agonist that activates GIRK channels without affecting neurotransmitter release.
Assuntos
Receptor 5-HT1A de Serotonina/metabolismo , Respiração/efeitos dos fármacos , Síndrome de Rett/tratamento farmacológico , Agonistas do Receptor 5-HT1 de Serotonina/farmacologia , Potenciais de Ação/efeitos dos fármacos , Animais , Apneia/metabolismo , Apneia/fisiopatologia , Encéfalo/metabolismo , Encéfalo/fisiopatologia , Modelos Animais de Doenças , Feminino , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/metabolismo , Ácido Glutâmico/metabolismo , Masculino , Proteína 2 de Ligação a Metil-CpG/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Piperidinas/farmacologia , Potássio/metabolismo , Pirimidinas/farmacologia , Núcleos da Rafe/efeitos dos fármacos , Núcleos da Rafe/metabolismo , Síndrome de Rett/metabolismo , Serotonina/metabolismo , Antagonistas do Receptor 5-HT1 de Serotonina/farmacologia , Transmissão Sináptica/efeitos dos fármacosRESUMO
The fetal respiratory response to acute hypoxia is characterized by depression, often to apnea. This study examined the effect of hypoxia on the electromyogram (EMG) of the thyroarytenoid (TA) muscle. Under anesthesia catheters were placed in the fetal sheep carotid artery, fourth cerebral ventricle, trachea and amniotic fluid and wires sewn into the diaphragm and TA muscle. During normoxic episodes of slow fetal breathing (<40 breaths per min) TA EMG activity was phasic beginning immediately after diaphragmatic EMG bursts and ending well before the next burst. This timing is consistent with the post-inspiratory (post-I) phase of the respiratory cycle. Lowering fetal arterial Pa O(2) from approximately 20mm Hg to approximately 13 mm Hg resulted in arrest of diaphragm EMG and tonic TA activity. Instillation of the (R,S)- -amino-3- hydroxy-5-methylisoxazole-4-proprionic acid (AMPA) ionotrophic glutamate receptor antagonist 2,3-dihydro-6-nitro-7-sulphamoyl-benzo(f) quinoxaline (NBQX) into the cerebrospinal fluid (CSF) of the fourth ventricle abolished tracheal pressure deflections and diaphragmatic EMG activity. Tonic TA activity, however, could still be evoked by hypoxia. These results indicate that fetal post-I motoneurons are not inhibited by moderate hypoxia and that their tonic activity may be due to a loss of inhibitory input.
Assuntos
Diafragma/fisiopatologia , Eletromiografia , Feto/fisiopatologia , Hipóxia/fisiopatologia , Músculos Laríngeos/fisiopatologia , Animais , Antagonistas de Aminoácidos Excitatórios/administração & dosagem , Injeções Intraventriculares , Quinoxalinas/administração & dosagem , Receptores de AMPA/antagonistas & inibidores , Mecânica Respiratória/efeitos dos fármacos , Taxa Respiratória/efeitos dos fármacos , OvinosRESUMO
Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X-linked gene methyl-CpG-binding protein 2 (MECP2) that encodes a DNA binding protein involved in gene silencing. Selective deletion of Mecp2 in post-mitotic neurons in mice results in a Rett-like phenotype characterized by disturbances in motor activity and body weight, suggesting that these symptoms are exclusively caused by neuronal deficiency. Included in the RTT phenotype are episodes of respiratory depression that follow hyperventilation. Here we show that the respiratory phenotype depends on the organ distribution of Mecp2 deficiency. Both female mice heterozygous for a null mutation in Mecp2 (Mecp2+/-) and those with selective deletion of the protein in neurons (Mecp2+/nestin-Cre lox), showed an initial response to hypoxia that exceeded that in wild type (WT). However, marked respiratory depression following hypoxic hyperventilation was only seen in Mecp2+/- animals. Addition of carbon dioxide to the hypoxic exposure eliminated the respiratory depression. Tidal volume and lung volume were larger in Mecp2+/- and respiratory depression was directly related to tidal volume. Taken together these results indicate that the depression is due to hypocapnia. Respiratory depression in this mouse model of Rett Syndrome is seen in with ubiquitous deficiency in Mecp2 but not when it is confined to neurons.