RESUMEN
Linalool and linalyl acetate are major components of lavender essential oil. These substances possess many biological activities, such as anti-inflammatory activity, analgesic and anxiolytic effects, and anticonvulsant properties, and they also induce modulation of neuronal activity in the autonomic nervous system. However, there are no reports of the direct effects of linalool on respiratory activity. In the present study, we analyzed the effects of linalool and linalyl acetate on central respiratory activity in the brainstem-spinal cord preparation isolated from newborn rats. Linalool dose-dependently decreased the rate of respiratory activity. This effect was reversed by bicuculline, suggesting that linalool enhanced inhibitory synaptic connections via GABAA receptors. In addition, linalool reduced the coefficient of variation of inspiratory burst intervals and thus could work to stabilize the respiratory rhythm. Linalyl acetate did not cause inhibitory effects as observed in linalool treatment. Linalool depressed burst activity of pre-inspiratory neurons in the medullary respiratory networks and increased the amplitude of inspiratory inhibitory postsynaptic potentials of pre-inspiratory neurons. We concluded that linalool caused inhibitory effects on respiratory rhythm generation mainly through activation of presynaptic GABAA receptors of pre-inspiratory neurons.
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Monoterpenos Acíclicos , Animales Recién Nacidos , Tronco Encefálico , Monoterpenos , Neuronas , Médula Espinal , Animales , Monoterpenos Acíclicos/farmacología , Médula Espinal/efectos de los fármacos , Médula Espinal/metabolismo , Ratas , Monoterpenos/farmacología , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Tronco Encefálico/efectos de los fármacos , Tronco Encefálico/fisiología , Receptores de GABA-A/metabolismo , Respiración/efectos de los fármacos , Bicuculina/farmacologíaRESUMEN
The lateral parafacial area (pFL) is a crucial region involved in respiratory control, particularly in generating active expiration through an expiratory oscillatory network. Active expiration involves rhythmic abdominal (ABD) muscle contractions during late-expiration, increasing ventilation during elevated respiratory demands. The precise anatomical location of the expiratory oscillator within the ventral medulla's rostro-caudal axis is debated. While some studies point to the caudal tip of the facial nucleus (VIIc) as the oscillator's core, others suggest more rostral areas. Our study employed bicuculline (a γ-aminobutyric acid type A [GABA-A] receptor antagonist) injections at various pFL sites (-0.2 mm to +0.8 mm from VIIc) to investigate the impact of GABAergic disinhibition on respiration. These injections consistently elicited ABD recruitment, but the response strength varied along the rostro-caudal zone. Remarkably, the most robust and enduring changes in tidal volume, minute ventilation, and combined respiratory responses occurred at more rostral pFL locations (+0.6/+0.8 mm from VIIc). Multivariate analysis of the respiratory cycle further differentiated between locations, revealing the core site for active expiration generation with this experimental approach. Our study advances our understanding of neural mechanisms governing active expiration and emphasizes the significance of investigating the rostral pFL region.
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Bicuculina , Espiración , Bicuculina/farmacología , Bicuculina/administración & dosificación , Animales , Espiración/fisiología , Masculino , Respiración/efectos de los fármacos , Bulbo Raquídeo/fisiología , Bulbo Raquídeo/efectos de los fármacos , Antagonistas de Receptores de GABA-A/farmacología , Antagonistas de Receptores de GABA-A/administración & dosificaciónRESUMEN
The reliable induction of long-term potentiation (LTP) in the dentate gyrus (DG) in vitro requires the blockade of the γ-aminobutyric acid A (GABAA) receptor. In these studies we examined the effectiveness of the specific GABAA receptor antagonist bicuculline methiodide (BMI) in facilitating LTP in the DG from hippocampal slices obtained from either C57Bl/6 mice or Sprague-Dawley rats, two species commonly used for electrophysiology. In the C57Bl/6 mice, maximal short-term potentiation and LTP in the DG were produced with a concentration of 5 µM BMI. In contrast, a concentration of 10 µM BMI was required to produce maximal short-term potentiation and LTP in the DG of Sprague-Dawley rats. These results reveal that there are species differences in the optimal amount of BMI required to produce robust and reliable LTP in the rodent DG in vitro and highlight the need to take consideration of the species being used when choosing concentrations of pharmacological agents to employ for electrophysiological use.NEW & NOTEWORTHY In this report we provide specific neurophysiological evidence for concentrations of GABAA antagonist required to study long-term potentiation in the medial perforant pathway of the dentate gyrus. Two commonly used species, Sprague-Dawley rats and C57Bl/6 mice, require different concentrations of bicuculline methiodide to induce optimal short-term and long-term potentiation.
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Bicuculina , Giro Dentado , Antagonistas de Receptores de GABA-A , Potenciación a Largo Plazo , Ratones Endogámicos C57BL , Ratas Sprague-Dawley , Animales , Potenciación a Largo Plazo/efectos de los fármacos , Potenciación a Largo Plazo/fisiología , Giro Dentado/efectos de los fármacos , Giro Dentado/fisiología , Bicuculina/farmacología , Bicuculina/análogos & derivados , Antagonistas de Receptores de GABA-A/farmacología , Ratones , Ratas , Masculino , Receptores de GABA-A/efectos de los fármacos , Receptores de GABA-A/metabolismo , Receptores de GABA-A/fisiología , Especificidad de la EspecieRESUMEN
BACKGROUND: Human induced pluripotent stem cell (hiPSC)- derived neurons offer the possibility of studying human-specific neuronal behaviors in physiologic and pathologic states in vitro. It is unclear whether cultured neurons can achieve the fundamental network behaviors required to process information in the brain. Investigating neuronal oscillations and their interactions, as occurs in cross-frequency coupling (CFC), addresses this question. NEW METHODS: We examined whether networks of two-dimensional (2D) cultured hiPSC-derived cortical neurons grown with hiPSC-derived astrocytes on microelectrode array plates recapitulate the CFC that is present in vivo. We employed the modulation index method for detecting phase-amplitude coupling (PAC) and used offline spike sorting to analyze the contribution of single neuron spiking to network behavior. RESULTS: We found that PAC is present, the degree of PAC is specific to network structure, and it is modulated by external stimulation with bicuculline administration. Modulation of PAC is not driven by single neurons, but by network-level interactions. COMPARISON WITH EXISTING METHODS: PAC has been demonstrated in multiple regions of the human cortex as well as in organoids. This is the first report of analysis demonstrating the presence of coupling in 2D cultures. CONCLUSION: CFC in the form of PAC analysis explores communication and integration between groups of neurons and dynamical changes across networks. In vitro PAC analysis has the potential to elucidate the underlying mechanisms as well as capture the effects of chemical, electrical, or ultrasound stimulation; providing insight into modulation of neural networks to treat nervous system disorders in vivo.
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Células Madre Pluripotentes Inducidas , Microelectrodos , Neuronas , Humanos , Neuronas/fisiología , Células Madre Pluripotentes Inducidas/fisiología , Células Madre Pluripotentes Inducidas/citología , Potenciales de Acción/fisiología , Células Cultivadas , Corteza Cerebral/fisiología , Corteza Cerebral/citología , Astrocitos/fisiología , Técnicas de Cultivo de Célula/métodos , Técnicas de Cultivo de Célula/instrumentación , Bicuculina/farmacología , Red Nerviosa/fisiologíaRESUMEN
Swallowing is induced by a central pattern generator in the nucleus tractus solitarius (NTS). We aimed to create a medullary slice preparation to elucidate the neural architecture of the central pattern generator of swallowing (Sw-CPG) and record its neural activities. Experiments were conducted on 2-day-old Sprague-Dawley rats (n = 46). The brainstem-spinal cord was transected at the pontomedullary and cervicothoracic junctions; the medulla was sliced transversely at thicknesses of 600, 700, or 800 µm. The rostral end of the slice was 100 µm rostral to the vagus nerve. We recorded hypoglossal nerve activity and electrically stimulated the vagus nerve or microinjected bicuculline methiodide (BIC) into the NTS. The 800-µm slices generated both rhythmic respiratory activity and electrically elicited neural activity. The 700-µm slices generated only respiratory activity, while the 600-µm slices did not generate any neural activity. BIC microinjection into the NTS in 800-µm slices resulted in the typical activity that closely resembled the swallowing activity reported in other experiments. This swallowing-like activity consistently lengthened the respiratory interval. Despite complete inhibition of respiratory activity, weak swallowing-like activity was observed under bath application of a non-NMDA receptor antagonist. Contrastingly, bath application of NMDA receptor antagonists resulted in a complete loss of swallowing-like activity and no change in respiratory activity. These results suggest that the 800-µm medullary slice preparation contains both afferent and efferent neural circuits and pattern generators of swallowing activity. Additionally, NMDA receptors may be necessary for generating swallowing activity. This medullary slice preparation can therefore elucidate Sw-CPG neural networks.
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Animales Recién Nacidos , Bicuculina , Generadores de Patrones Centrales , Deglución , Nervio Hipogloso , Bulbo Raquídeo , Ratas Sprague-Dawley , Nervio Vago , Animales , Deglución/fisiología , Deglución/efectos de los fármacos , Bulbo Raquídeo/fisiología , Bulbo Raquídeo/efectos de los fármacos , Bicuculina/farmacología , Bicuculina/análogos & derivados , Ratas , Nervio Vago/fisiología , Nervio Vago/efectos de los fármacos , Generadores de Patrones Centrales/fisiología , Generadores de Patrones Centrales/efectos de los fármacos , Nervio Hipogloso/fisiología , Nervio Hipogloso/efectos de los fármacos , Estimulación Eléctrica , Núcleo Solitario/efectos de los fármacos , Núcleo Solitario/fisiologíaRESUMEN
Epilepsy designates a group of chronic brain disorders, characterized by the recurrence of hypersynchronous, repetitive activity, of neuronal clusters. Epileptic seizures are the hallmark of epilepsy. The primary goal of epilepsy treatment is to eliminate seizures with minimal side effects. Nevertheless, approximately 30% of patients do not respond to the available drugs. An imbalance between excitatory/inhibitory neurotransmission, that leads to excitotoxicity, seizures, and cell death, has been proposed as an important mechanism regarding epileptogenesis. Recently, it has been shown that microreactors composed of platinum nanoparticles (Pt-NP) and glutamate dehydrogenase possess in vitro and in vivo activity against excitotoxicity. This study investigates the in vivo effects of these microreactors in an animal model of epilepsy induced by the administration of the GABAergic antagonist bicuculline. Male Wistar rats were administered intracerebroventricularly (i.c.v.) with the microreactors or saline and, five days later, injected with bicuculline or saline. Seizure severity was evaluated in an open field. Thirty min after behavioral measurements, animals were euthanized, and their brains processed for neurodegeneration evaluation and for neurogenesis. Treatment with the microreactors significantly increased the time taken for the onset of seizures and for the first tonic-clonic seizure, when compared to the bicuculline group that did not receive the microreactor. The administration of the microreactors also increased the time spent in total exploration and grooming. Treatment with the microreactors decreased bicuculline-induced neurodegeneration and increased neurogenesis in the dorsal and ventral hippocampus. These observations suggest that treatment with Pt-NP-based microreactors attenuates the behavioral and neurobiological consequences of epileptiform seizure activity.
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Epilepsia , Nanopartículas del Metal , Fármacos Neuroprotectores , Humanos , Ratas , Animales , Masculino , Bicuculina/farmacología , Platino (Metal)/efectos adversos , Ratas Wistar , Convulsiones/inducido químicamente , Convulsiones/tratamiento farmacológicoRESUMEN
Allopregnanolone (ALLO) is a known neurosteroid and a progesterone metabolite synthesized in the ovary, CNS, PNS, adrenals and placenta. Its role in the neuroendocrine control of ovarian physiology has been studied, but its in situ ovarian effects are still largely unknown. The aims of this work were to characterize the effects of intrabursal ALLO administration on different ovarian parameters, and the probable mechanism of action. ALLO administration increased serum progesterone concentration and ovarian 3ß-HSD2 while decreasing 20α-HSD mRNA expression. ALLO increased the number of atretic follicles and the number of positive TUNEL granulosa and theca cells, while decreasing positive PCNA immunostaining. On the other hand, there was an increase in corpora lutea diameter and PCNA immunostaining, whereas the count of TUNEL-positive luteal cells decreased. Ovarian angiogenesis and the immunohistochemical expression of GABAA receptor increased after ALLO treatment. To evaluate if the ovarian GABAA receptor was involved in these effects, we conducted a functional experiment with a specific antagonist, bicuculline. The administration of bicuculline restored the number of atretic follicles and the diameter of corpora lutea to normal values. These results show the actions of ALLO on the ovarian physiology of the female rat during the follicular phase, some of them through the GABAA receptor. Intrabursal ALLO administration alters several processes of the ovarian morpho-physiology of the female rat, related to fertility and oocyte quality.
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Pregnanolona , Progesterona , Embarazo , Femenino , Ratas , Animales , Pregnanolona/farmacología , Progesterona/farmacología , Antígeno Nuclear de Célula en Proliferación , Bicuculina/farmacología , Receptores de GABA-A , Cuerpo LúteoRESUMEN
Imaging hemodynamic responses to interictal spikes holds promise for presurgical epilepsy evaluations. Understanding the hemodynamic response function is crucial for accurate interpretation. Prior interictal neurovascular coupling data primarily come from anesthetized animals, impacting reliability. We simultaneously monitored calcium fluctuations in excitatory neurons, hemodynamics, and local field potentials (LFP) during bicuculline-induced interictal events in both isoflurane-anesthetized and awake mice. Isoflurane significantly affected LFP amplitude but had little impact on the amplitude and area of the calcium signal. Anesthesia also dramatically blunted the amplitude and latency of the hemodynamic response, although not its area of spread. Cerebral blood volume change provided the best spatial estimation of excitatory neuronal activity in both states. Targeted silencing of the thalamus in awake mice failed to recapitulate the impact of anesthesia on hemodynamic responses suggesting that isoflurane's interruption of the thalamocortical loop did not contribute either to the dissociation between the LFP and the calcium signal nor to the alterations in interictal neurovascular coupling. The blood volume increase associated with interictal spikes represents a promising mapping signal in both the awake and anesthetized states.
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Hemodinámica , Isoflurano , Neuronas , Vigilia , Animales , Ratones , Vigilia/efectos de los fármacos , Vigilia/fisiología , Hemodinámica/efectos de los fármacos , Neuronas/efectos de los fármacos , Isoflurano/farmacología , Anestesia , Masculino , Circulación Cerebrovascular/efectos de los fármacos , Circulación Cerebrovascular/fisiología , Ratones Endogámicos C57BL , Bicuculina/farmacología , Acoplamiento Neurovascular/efectos de los fármacos , Acoplamiento Neurovascular/fisiologíaRESUMEN
Gamma-aminobutyric acid (GABA) disinhibition in medial hypothalamus (MH) nuclei of rats elicits some defensive reactions that are considered panic attack-like behaviours. Recent evidence showed that the norepinephrine-mediated system modulates fear-related defensive behaviours organised by MH neurons at least in part via noradrenergic receptors recruitment on midbrain tegmentum. However, it is unknown whether noradrenergic receptors of the MH also modulate the panic attack-like reactions. The aim of this work was to investigate the distribution of noradrenergic receptors in MH, and the effects of either α1-, α2- or ß-noradrenergic receptors blockade in the MH on defensive behaviours elaborated by hypothalamic nuclei. Defensive behaviours were evaluated after the microinjection of the selective GABAA receptor antagonist bicuculline into the MH that was preceded by microinjection of either WB4101, RX821002, propranolol (α1-, α2- and ß-noradrenergic receptor selective antagonists, respectively), or physiological saline into the MH of male Wistar rats. The α1-, α2- and ß-noradrenergic receptors were found in neuronal perikarya of all MH nuclei, and the α2-noradrenergic receptor were also found on glial cells mainly situated in the ventrolateral division of the ventromedial hypothalamic nucleus. The α1- and ß-noradrenergic receptors blockade in the MH decreased defensive attention and escape reactions elicited by the intra-MH microinjections of bicuculline. These findings suggest that, despite the profuse distributions of α1-, α2- and ß-noradrenergic receptors in the MH, both α1- and ß-noradrenergic receptor- rather than α2-noradrenergic receptor-signalling in MH are critical for the neuromodulation of panic-like behaviour.
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Trastorno de Pánico , Ratas , Masculino , Animales , Núcleo Hipotalámico Ventromedial , Bicuculina/farmacología , Ratas Wistar , Transmisión Sináptica , MicroinyeccionesRESUMEN
We investigated the effects of histamine and GABA A receptor agents on pain and depression-like behaviors and their interaction using a tail-flick test and the forced swimming test (FST) in male mice. Our data revealed that intraperitoneal administration of muscimol (0.12 and 0.25â mg/kg) increased the percentage of maximum possible effect (%MPE) and area under the curve (AUC) of %MPE, indicating an antinociceptive response. Intraperitoneal injection of bicuculline (0.5 and 1â mg/kg) decreased %MPE and AUC of %MPE, suggesting hyperalgesia. Moreover, muscimol by reducing the immobility time of the FST elicited an antidepressant-like response but bicuculline by enhancing the immobility time of the FST caused a depressant-like response. Intracerebroventricular (i.c.v.) microinjection of histamine (5â µg/mouse) enhanced %MPE and AUC of %MPE. i.c.v. infusion of histamine (2.5 and 5â µg/mouse) decreased immobility time in the FST. Co-administration of different doses of histamine along with a sub-threshold dose of muscimol potentiated antinociceptive and antidepressant-like responses produced by histamine. Cotreatment of different doses of histamine plus a noneffective dose of bicuculline reversed antinociception and antidepressant-like effects elicited by histamine. Cotreatment of histamine, muscimol, and bicuculline reversed antinociceptive and antidepressant-like behaviors induced by the drugs. The results demonstrated additive antinociceptive and antidepressant-like effects between histamine and muscimol in mice. In conclusion, our results indicated an interaction between the histaminergic and GABAergic systems in the modulation of pain and depression-like behaviors.
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Antidepresivos , Histamina , Ratones , Masculino , Animales , Muscimol/farmacología , Histamina/farmacología , Bicuculina/farmacología , Antidepresivos/farmacología , Natación , Analgésicos/farmacología , Dolor/tratamiento farmacológicoRESUMEN
Goal of this work is to show how the developmental conditions of in vitro neuronal networks influence the effect of drug delivery. The proposed experimental neuronal model consists of dissociated cortical neurons plated to Micro-Electrode Arrays (MEAs) and grown according to different conditions (i.e., by varying both the adopted culture medium and the number of days needed to let the network grow before performing the chemical modulation). We delivered rising amount of bicuculline (BIC), a competitive antagonist of GABAA receptors, and we computed the firing rate dose-response curve for each culture. We found that networks matured in BrainPhys for 18 days in vitro exhibited a decreasing firing trend as a function of the BIC concentration, quantified by an average IC50 (i.e., half maximal inhibitory concentration) of 4.64 ± 4.02 µM. On the other hand, both cultures grown in the same medium for 11 days, and ones matured in Neurobasal for 18 days displayed an increasing firing rate when rising amounts of BIC were delivered, characterized by average EC50 values (i.e., half maximal excitatory concentration) of 0.24 ± 0.05 µM and 0.59 ± 0.46 µM, respectively.Clinical Relevance- This research proves the relevance of the experimental factors that can influence the network development as key variables when developing a neuronal model to conduct drug delivery in vitro, simulating the in vivo environment. Our findings suggest that not considering the consequences of the chosen growing conditions when performing in vitro pharmacological studies could lead to incomplete predictions of the chemically induced alterations.
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Neuronas , Bicuculina/farmacología , Neuronas/fisiología , ElectrodosRESUMEN
Fossorial Damaraland mole-rats (Fukomys damarensis) mount a robust hypoxic metabolic response (HMR) but a blunted hypoxic ventilatory response (HVR) to acute hypoxia. Although these reflex physiological responses have been described previously, the underlying signalling pathways are entirely unknown. Of particular interest are contributions from γ-aminobutyric acid (GABA), which is the primary inhibitory neurotransmitter in the nervous system of most adult mammals, and adenosine, the accumulation of which increases during hypoxia as a breakdown product of ATP. Therefore, we hypothesized that GABAergic and/or adenosinergic signalling contributes to the blunted HVR and robust HMR in Damaraland mole-rats. To test this hypothesis, we injected adult animals with saline alone (controls), or 100â mgâ kg-1 aminophylline or 1â mgâ kg-1 bicuculline, to block adenosine or GABAA receptors, respectively. We then used respirometry, plethysmography and thermal RFID probes to non-invasively measure metabolic, ventilator and thermoregulatory responses, respectively, to acute hypoxia (1â h in 5 or 7% O2) in awake and freely behaving animals. We found that bicuculline had relatively minor effects on metabolism and thermoregulation but sensitized ventilation such that the HVR became manifest at 7% instead of 5% O2 and was greater in magnitude. Aminophylline increased metabolic rate, ventilation and body temperature in normoxia, and augmented the HMR and HVR. Taken together, these findings indicate that adenosinergic and GABAergic signalling play important roles in mediating the robust HMR and blunted HVR in Damaraland mole-rats.
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Adenosina , Aminofilina , Animales , Bicuculina/farmacología , Adenosina/farmacología , Ratas Topo/fisiología , Hipoxia/metabolismo , Ácido gamma-AminobutíricoRESUMEN
The neurotransmitter γ-aminobutyric acid (GABA) drives critical inhibitory processes in and beyond the nervous system, partly via ionotropic type-A receptors (GABAARs). Pharmacological properties of ρ-type GABAARs are particularly distinctive, yet the structural basis for their specialization remains unclear. Here, we present cryo-EM structures of a lipid-embedded human ρ1 GABAAR, including a partial intracellular domain, under apo, inhibited, and desensitized conditions. An apparent resting state, determined first in the absence of modulators, was recapitulated with the specific inhibitor (1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid and blocker picrotoxin and provided a rationale for bicuculline insensitivity. Comparative structures, mutant recordings, and molecular simulations with and without GABA further explained the sensitized but slower activation of ρ1 relative to canonical subtypes. Combining GABA with picrotoxin also captured an apparent uncoupled intermediate state. This work reveals structural mechanisms of gating and modulation with applications to ρ-specific pharmaceutical design and to our biophysical understanding of ligand-gated ion channels.
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Receptores de GABA-A , Ácido gamma-Aminobutírico , Humanos , Receptores de GABA-A/metabolismo , Picrotoxina/farmacología , Ligandos , Ácido gamma-Aminobutírico/metabolismo , Bicuculina/farmacología , Sitios de UniónRESUMEN
Bicuculline and saclofen were microinjected into the rostral (rNTS) and caudal nucleus of the solitary tract (cNTS) in 17 anesthetized cats. Electromyograms (EMGs) of the diaphragm (DIA) and abdominal muscles (ABD), esophageal pressures (EP), and blood pressure were recorded and analyzed. Bilateral microinjections of 1 mM bicuculline in the rNTS significantly reduced the number of coughs (CN), amplitudes of DIA and ABD EMG, inspiratory and expiratory EP, and prolonged the duration of the cough expiratory phase (CTE) as well as the total cough cycle duration (CTtot). Bilateral microinjections of 2 mM saclofen reduced only cough expiratory efforts. Bilateral microinjection of bicuculline in the cNTS significantly reduced CN and amplitudes of ABD EMG and elongated CTE and CTtot. Bilateral microinjections of saclofen in cNTS had no significant effect on analyzed cough parameters. Our results confirm a different GABAergic inhibitory system in the rNTS and cNTS acting on mechanically induced cough in cats.
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Tos , Núcleo Solitario , Gatos , Animales , Tos/tratamiento farmacológico , Bicuculina/farmacología , Antagonistas de Receptores de GABA-B/farmacología , Antagonistas de Receptores de GABA-B/uso terapéutico , Baclofeno/farmacología , MicroinyeccionesRESUMEN
Abnormal depolarization of neuronal membranes called paroxysmal depolarization shift (PDS) represents a cellular correlate of interictal spikes. The mechanisms underlying the generation of PDSs or PDS clusters remain obscure. This study aimed to investigate the role of ionotropic glutamate receptors (iGluRs) in the generation of PDS and dependence of the PDS pattern on neuronal membrane potential. We have shown that significant depolarization or hyperpolarization (by more than ±50 mV) of a single neuron does not change the number of individual PDSs in the cluster, indicating the involvement of an external stimulus in PDS induction. Based on this data, we have suggested reliable protocols for stimulating single PDS or PDS clusters. Furthermore, we have found that AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptors are necessary for PDS generation since AMPAR antagonist NBQX completely suppresses bicuculline-induced paroxysmal activity. In turn, antagonists of NMDA (N-methyl-D-aspartate) and kainate receptors (D-AP5 and UBP310, respectively) caused a decrease in the amplitude of the first action potential in PDSs and in the amplitude of the oscillations of intracellular Ca2+ concentration occurring alongside the PDS cluster generation. The effects of the NMDAR (NMDA receptor) and KAR (kainate receptor) antagonists indicate that these receptors are involved only in the modulation of paroxysmal activity. We have also shown that agonists of some Gi-coupled receptors, such as A1 adenosine (A1Rs) or cannabinoid receptors (CBRs) (N6-cyclohexyladenosine and WIN 55,212-2, respectively), completely suppressed PDS generation, while the A1R agonist even prevented it. We hypothesized that the dynamics of extracellular glutamate concentration govern paroxysmal activity. Fine-tuning of neuronal activity via action on Gi-coupled receptors or iGluRs paves the way for the development of new approaches for epilepsy pharmacotherapy.
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Hipocampo , Receptores de N-Metil-D-Aspartato , Ratas , Animales , Bicuculina/farmacología , Neuronas , Potenciales de Acción , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiónico/farmacologíaRESUMEN
Although ethanol administration produces a range of physiological effects, the rewarding aspect associated with its consumption is a major contributory factor to its abuse liability. Recently, lateral habenula (LHb) has been shown to be engaged by both rewarding and aversive stimuli. Its major glutamatergic output, the fasciculus retroflexus, projects to the rostromedial tegmental nucleus (RMTg) and controls the activity of the ventral tegmental area (VTA) dopaminergic system to promote reward circuitry. While several attempts have been made to understand the relationship between LHb and addiction, there is still a lack of knowledge in relation to ethanol addiction. In the present study, by pharmacologically exacerbating or inhibiting the LHb or RMTg neuronal activity during a post-conditioning test, we investigated the role of LHb-RMTg fasciculus retroflexus in ethanol-induced reward behavior using the conditioned place preference (CPP) test. We found that activation of LHb glutamatergic system by intra-LHb administration of l-trans-2,4-pyrrolidine dicarboxylate (PDC) (glutamate transporter inhibitor) significantly decreased CPP score; on the contrary, lamotrigine (inhibits glutamate release) significantly increased CPP score and showed a rewarding effect in CPP. Instead, intra-RMTg administration of muscimol (GABAA receptor agonist) significantly increased CPP score, whereas bicuculline (GABAA antagonist) treatment decreased CPP score. In immunohistochemistry, we found that PDC administration significantly decreased, whereas lamotrigine treatment significantly increased tyrosine hydroxylase immunoreactivity (TH-ir) in VTA and nucleus accumbens (NAc). Furthermore, while intra-RMTg administration of muscimol increased, the bicuculline treatment significantly decreased the TH-ir in VTA and NAc. Together, our behavioral and immunohistochemical results signify the role of LHb and RMTg in the expression of ethanol-conditioned reward behavior.
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Habénula , Habénula/metabolismo , Bicuculina/farmacología , Bicuculina/metabolismo , Lamotrigina/metabolismo , Muscimol/farmacología , Muscimol/metabolismo , Área Tegmental Ventral/fisiología , Etanol/farmacología , Etanol/metabolismoRESUMEN
OBJECTIVE: Focal epilepsy is thought to be a network disease, in which epileptiform activity can spread noncontiguously through the brain via highly interconnected nodes, or hubs, within existing networks. Animal models confirming this hypothesis are scarce, and our understanding of how distant nodes are recruited is also lacking. Whether interictal spikes (IISs) also create and reverberate through a network is not well understood. METHODS: We injected bicuculline into the S1 barrel cortex and employed multisite local field potential and Thy-1 and parvalbumin (PV) cell mesoscopic calcium imaging during IISs to monitor excitatory and inhibitory cells in two monosynaptically connected nodes and one disynaptically connected node: ipsilateral secondary motor area (iM2), contralateral S1 (cS1), and contralateral secondary motor area (cM2). Node participation was analyzed with spike-triggered coactivity maps. Experiments were repeated with 4-aminopyridine as an epileptic agent. RESULTS: We found that each IIS reverberated throughout the network, differentially recruiting both excitatory and inhibitory cells in all connected nodes. The strongest response was found in iM2. Paradoxically, node cM2, which was connected disynaptically to the focus, was recruited more intensely than node cS1, which was connected monosynaptically. The explanation for this effect could be found in node-specific excitatory/inhibitory (E/I) balance, as cS1 demonstrated greater PV inhibitory cell activation compared with cM2, where Thy-1 excitatory cells were more heavily recruited. SIGNIFICANCE: Our data show that IISs spread noncontiguously by exploiting fiber pathways that connect nodes in a distributed network and that E/I balance plays a critical role in node recruitment. This multinodal IIS network model can be used to investigate cell-specific dynamics in the spatial propagation of epileptiform activity.
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Epilepsia , Animales , Encéfalo , Mapeo Encefálico , Bicuculina/farmacología , 4-AminopiridinaRESUMEN
The relationship between serotonin dysfunction and schizophrenia commenced with the discovery of the effects of lysergic acid diethylamide (LSD) that has high affinity for 5-HT2A receptors. Activation of these receptors produces perceptual and behavioural changes such as illusions, visual hallucinations and locomotor hyperactivity. Using prepulse inhibition (PPI) of the acoustic startle, which is impaired in schizophrenia,we aimed to investigate:i) the existence of a direct and potentially inhibitory neural pathway between the inferior colliculus (IC) and the pedunculopontine tegmental nucleus (PPTg) involved in the mediation of PPI responses by a neural tract tracing procedure;ii) if the microinjection of the 5-HT2A receptors agonist DOI in IC would activate neurons in this structure and in the PPTg by a c-Fos protein immunohistochemistry study;iii) whether the deficits in PPI responses, observed after the administration of DOI in the IC, could be prevented by the concomitant microinjection of the GABAA receptor antagonist bicuculline in the PPTg.Male Wistar rats were used in this study. An IC-PPTg reciprocated neuronal pathway was identified by neurotracing. The number of c-Fos labelled cells was lower in the DOI group in IC and PPTg, suggesting that this decrease could be due to the high levels of GABA in both structures. The concomitant microinjections of bicuculline in PPTg and DOI in IC prevented the PPI deficit observed after the IC microinjection of DOI. Our findings suggest that IC 5-HT2A receptors may be at least partially involved in the regulation of inhibitory pathways mediating PPI response in IC and PPTg structures.
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Colículos Inferiores , Núcleo Tegmental Pedunculopontino , Ratas , Animales , Masculino , Inhibición Prepulso/fisiología , Reflejo de Sobresalto/fisiología , Receptores de GABA-A , Receptor de Serotonina 5-HT2A , Bicuculina/farmacología , Serotonina/farmacología , Ratas WistarRESUMEN
Seizure-like burst activities are induced by blockade of GABAA and/or glycine receptors in various spinal ventral roots of brainstem-spinal cord preparation from neonatal rodents. We found that this is not applicable to the phrenic nerve and that a new inhibitory descending pathway may suppress seizure-like activity in the phrenic nerve. Experiments were performed in brainstem-spinal cord preparation from newborn rats (age: 0-1 day). Left phrenic nerve and right C4 activities were recorded simultaneously. When GABAA and glycine receptors were blocked by 10 µM bicuculline and 10 µM strychnine (Bic+Str), seizure-like burst activities appeared in the fourth cervical ventral root (C4) but not the phrenic nerve. After making a transverse section at C1, the inspiratory burst activity disappeared from both C4 and the phrenic nerve, whereas seizure-like activity appeared in both nerves. We hypothesized that inhibitory descending pathways other than those via GABAA and/or glycine receptors (from the medulla to the spinal cord) work to avoid disturbance of regular respiratory-related diaphragm contraction by seizure-like activity. We found that cannabinoid receptor antagonist, AM251 was effective for the induction of seizure-like activity by Bic+Str in the phrenic nerve in brainstem-spinal cord preparation. Cannabinoid receptors may be involved in this descending inhibitory system.
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Receptores de Glicina , Médula Espinal , Animales , Ratas , Animales Recién Nacidos , Receptores de Cannabinoides , Bicuculina/farmacología , Estricnina/farmacología , Convulsiones/tratamiento farmacológico , Nervio Frénico/fisiologíaRESUMEN
This study investigated the interactive effect of glucocorticoid and Gamma-aminobutyric acid (GABA) receptors in the Infralimbic (IL) cortex on fear extinction in rats' auditory fear conditioning task (AFC). Animals received 3 conditioning trial tones (conditioned stimulus, 30 s, 4 kHz, 80 dB) co-terminated with a footshock (unconditioned stimulus, 0.8 mA, 1 s). Extinction testing was conducted over 3 days (Ext 1-3) after conditioning. Intra-IL injection of corticosterone (CORT, 20 ng/0.3 µl/side) was performed 15 min before the first extinction trial (Ext 1) which attenuated auditory fear expression in subsequent extinction trials (Ext 1-3), demonstrating fear memory extinction enhancement. Co-injection of the GABAA agonist muscimol (250 ng/0.3 µl/side) or the GABAB agonist baclofen (250 ng/0.3 µl/side) 15 min before corticosterone, did not significantly affect the facilitative effects of corticosterone on fear extinction. However, co-injection of the GABAA antagonist bicuculline (BIC, 100 ng/0.3 µl/side) or the GABAB antagonist CGP35348 (CGP, 100 ng/0.3 µl/side) 15 min before corticosterone, blocked the facilitative effects of corticosterone on fear extinction. Moreover, extracellular signal-regulated kinase (ERK) and cAMP response element-binding (CREB) in the IL were examined by Western blotting analysis after the first extinction trial (Ext 1) in some groups. Intra-IL injection of corticosterone increased the ERK activity but not CREB. Co-injection of the bicuculline or CGP35348 blocked the enhancing effect of corticosterone on ERK expression in the IL. Glucocorticoid receptors (GRs) activation in the IL cortex by corticosterone increased ERK activity and facilitated fear extinction. GABAA or GABAB antagonists decreased ERK activity and inhibited corticosterone's effect. GRs and GABA receptors in the IL cortex jointly modulate the fear extinction processes via the ERK pathway. This pre-clinical animal study may highlight GRs and GABA interactions in the IL cortex modulating fear memory processes in fear-related disorders such as post-traumatic stress disorder (PTSD).