Mirko Krolo

Modulation of the synaptic drive to respiratory premotor and motor neurons.

The characteristics of GABAergic inhibitory modulation of respiratory bulbospinal neuronal activity and short-term potentiation (STP) of phrenic motoneuronal activity were studied. Extracellular unit recording and picoejection techniques in anesthetized dogs showed that both the spontaneous rhythmic and reflexly induced discharge patterns of inspiratory (I) and expiratory (E) premotor neurons were proportionately amplified by the localized application of picomole amounts of bicuculline (Bic), a competitive GABAA antagonist. Intracellular recording and paired-pulse stimulation techniques in anesthetized rats demonstrated an STP of phrenic motor output that appears to be mediated by NMDA receptors and is associated with facilitation of EPSPs and prolonged depolarization of individual phrenic motoneurons. We speculate that both GABAergic gain modulation of premotor neuronal activity and NMDA-mediated STP of phrenic activity may be neural substrates which are involved with the optimization of respiratory and non-respiratory behaviors, via adaptive and/or differential control of breathing.

Effects of halothane on synaptic neurotransmission to medullary expiratory neurons in the ventral respiratory group of dogs.

BACKGROUND The activity of canine expiratory neurons is primarily dependent on N-methyl-D-aspartic acid (NMDA)-receptor mediated excitatory chemodrive inputs and a powerful inhibitory gain modulatory mechanism mediated via gamma-aminobutyric acidA (GABA(A)) receptors. We examined whether the depressant effect of halothane on expiratory neuronal activity is primarily caused by a reduction in glutamatergic excitation or a potentiation of the inhibitory mechanism. METHODS Experiments were performed in halothane-anesthetized, vagotomized, paralyzed, and mechanically ventilated dogs during hypercapnic hyperoxia. The effect of a halothane dose increase from one minimum alveolar concentration (MAC) to 2 MAC on extracellularly recorded expiratory neuronal activity was studied before and during complete GABA(A) receptor blockade by localized picoejection of bicuculline close to the neuron. Complete blockade of the inhibitory mechanism allowed differentiation between the effects of halothane on overall NMDA-mediated excitation and on GABA(A)-mediated inhibition. RESULTS The spontaneous activity of 12 expiratory neurons was significantly depressed (18.1%) by the 1-MAC halothane dose increase. Overall glutamatergic excitation was depressed 38.3+/-12.3% (mean +/- SD) by the 1-MAC halothane increase. The prevailing GABA(A)ergic attenuation of neuronal output decreased significantly from 49.5+/-10 to 32.0+/-10.4%. Thus overall inhibition was reduced by halothane by 33.5+/-17.2%. CONCLUSIONS These results suggest that the depressive effect of a 1-MAC halothane dose increase on expiratory neuronal activity in our in vivo preparation with an intact neural network was mainly caused by a reduction of synaptic excitatory mechanisms and not an enhancement of synaptic inhibitory mechanisms.

Effects of halothane on excitatory neurotransmission to medullary expiratory neurons in a decerebrate dog model.

BackgroundThe activity of canine expiratory (E) neurons in the caudal ventral respiratory group is primarily dependent on N-methyl-d-aspartic acid (NMDA) receptor–mediated excitatory chemodrive inputs and modulated by an inhibitory mechanism mediated via &ggr;-aminobutyric acidA (GABAA) receptors. In an intact canine preparation, halothane depressed the activity of these neurons mainly by reduction in overall glutamatergic excitation. A new decerebrate preparation allows comparison of the effects of halothane on these synaptic mechanisms with an anesthetic-free baseline state. MethodsTwo separate studies were performed in decerebrate, vagotomized, paralyzed, mechanically ventilated dogs during hypercapnic hyperoxia. In study 1, the effect of 1 minimum alveolar concentration (MAC) halothane on extracellularly recorded E neuronal activity was studied before and during complete GABAA receptor blockade by localized pressure ejection of bicuculline. Complete blockade of the inhibitory mechanism allowed differentiation between the effects of halothane on overall GABAA-mediated inhibition and on overall NMDA receptor–mediated excitation. In study 2, the effect of 1 MAC halothane on the dose response of neurons to localized picoejection of the glutamate agonist NMDA was used to estimate halothane effect on postsynaptic glutamatergic excitatory neurotransmission. ResultsIn study 1, the spontaneous activity of 14 E neurons was depressed 38.6 ± 20.6% (mean ± SD) by 1 MAC halothane. Overall excitation was depressed 31.5 ± 15.5%. The GABAergic inhibition showed a 11.7 ± 18.3% enhancement during halothane. In study 2, the spontaneous activity of 13 E neurons was again significantly depressed by 1 MAC halothane (27.9 ± 10.6%), but the postsynaptic response of the neurons to exogenous NMDA was not significantly depressed by halothane (3.3 ± 38.4%). ConclusionsTogether these results suggest that in our E neuron paradigm, halothane exerted its depressive effect mainly via reduction of glutamatergic presynaptic mechanisms.

Sevoflurane Enhances γ-Aminobutyric Acid Type A Receptor Function and Overall Inhibition of Inspiratory Premotor Neurons in a Decerebrate Dog Model

Background:Inspiratory premotor neurons in the caudal ventral medulla relay excitatory drive to phrenic and inspiratory intercostal motoneurons in the spinal cord. These neurons are subject to tonic &ggr;-aminobutyric acid type A (GABAA)ergic inhibition. In a previous study, 1 minimum alveolar concentration (MAC) sevoflurane depressed overall glutamatergic excitatory drive and enhanced overall GABAAergic inhibitory drive to the neurons. This study investigated in further detail the effects of sevoflurane on GABAAergic inhibition by examining postsynaptic GABAA receptor activity in these neurons. Methods:Studies were performed in decerebrate, vagotomized, paralyzed, and mechanically ventilated dogs during hypercapnic hyperoxia. The effect of 1 MAC sevoflurane on extracellularly recorded neuronal activity was measured during localized picoejection of the GABAA receptor antagonist bicuculline and the GABAA agonist muscimol. Complete blockade of GABAAergic inhibition by bicuculline allowed estimation of the prevailing overall inhibition of the neuron. The neuronal response to muscimol was used to assess the anesthetic effect on the postsynaptic GABAA receptor function. Results:One MAC sevoflurane depressed the spontaneous activity of 21 inspiratory premotor neurons by (mean ± SD) 32.6 ± 20.5% (P < 0.001). Overall excitatory drive was depressed 17.9 ± 19.8% (P < 0.01). Overall GABAAergic inhibition was enhanced by 18.5 ± 18.2% (P < 0.001), and the postsynaptic GABAA receptor function was increased by 184.4 ± 121.8% (n = 20; P < 0.001). Conclusion:One MAC sevoflurane greatly enhanced GABAA receptor function on inspiratory premotor neurons and increased overall synaptic inhibition but to a smaller extent, indicating that the presynaptic inhibitory input was also reduced. Therefore, the anesthetic depression of spontaneous inspiratory premotor neuronal activity by 1 MAC sevoflurane in vivo is due to a combined effect on the two major ionotropic synaptic neurotransmitter systems with a decrease in overall glutamatergic excitation and a strong enhancement of postsynaptic GABAA receptor function.

Dose-dependent effects of halothane on the phrenic nerve responses to acute hypoxia in vagotomized dogs.

Background: Previous studies in dogs and humans suggest that the carotid body chemoreceptor response to hypoxia is selectively impaired by halothane. The present studies in an open‐loop canine preparation were performed to better delineate the effects of anesthetic concentrations of halothane on the carotid body chemoreceptor‐mediated phrenic nerve response to an acute hypoxic stimulus. Methods: Three protocols were performed to study the effects of halothane anesthesia on the phrenic nerve response to 1 min of isocapnic hypoxia (partial pressure of oxygen [PaO2] at peak hypoxia, 35–38 mmHg) in unpremedicated, anesthetized, paralyzed, vagotomized dogs during constant mechanical ventilation. In protocol 1, the dose‐dependent effects of halothane from 0.5–2.0 minimum alveolar concentration (MAC) on the hypoxic response during moderate hypercapnia (partial pressure of carbon dioxide [PaCO2], 60–65 mmHg) were studied in 10 animals. In protocol 2, the hypoxic responses at 1 MAC halothane near normocapnia (PaCO2, 40–45 mmHg) and during moderate hypercapnia were compared in an additional four animals. In protocol 3, the hypoxic response of 4 of 10 dogs from protocol 1 was also studied under sodium thiopental (STP) anesthesia after they completed protocol 1. Results: Protocol 1: Peak phrenic nerve activity (PPA) increased significantly during the hypoxic runs compared with the isocapnic hyperoxic controls at all halothane doses. The phrenic nerve response to the hypoxic stimulus was present even at the 2 MAC dose. Protocol 2: The net hypoxic responses for the two carbon dioxide background levels at 1 MAC were not significantly. Protocol 3: The net hypoxic response of PPA for the STP anesthetic was not significantly different from the 1 MAC halothane dose. Bilateral carotid sinus denervation abolished the PPA response to hypoxia. Conclusions: The phrenic nerve response to an acute, moderately severe isocapnic hypoxic stimulus is dose‐dependently depressed but not abolished by surgical doses of halothane. This analysis does not suggest a selective depression of the carotid body chemoreceptor response by halothane. The observed hypoxic phrenic response was mediated by the carotid body chemoreceptors in vagotomized dogs because bilateral carotid sinus denervation abolished all increases in PPA.

Sevoflurane Depresses Glutamatergic Neurotransmission to Brainstem Inspiratory Premotor Neurons but Not Postsynaptic Receptor Function in a Decerebrate Dog Model

Background:Inspiratory bulbospinal neurons in the caudal ventral medulla are premotor neurons that drive motoneurons, which innervate pump muscles such as the diaphragm and external intercostals. Excitatory drive to these neurons is mediated by N-methyl-d-aspartate (NMDA) receptors and &agr;-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors and is modulated by an inhibitory &ggr;-aminobutyric acid type A (GABAA)ergic input. The authors investigated the effect of sevoflurane on these synaptic mechanisms in decerebrate dogs. Methods:Studies were performed in decerebrate, vagotomized, paralyzed, and mechanically ventilated dogs during hypercapnic hyperoxia. The effect of 1 minimum alveolar concentration sevoflurane on extracellularly recorded activity of single neurons was measured during localized picoejection of the GABAA receptor blocker bicuculline and the glutamate agonists AMPA and NMDA. Complete blockade of the GABAAergic mechanism by bicuculline allowed differentiation between the effects of sevoflurane on overall GABAAergic inhibition and on overall glutamatergic excitation. The neuronal responses to exogenous AMPA and NMDA were used to estimate the anesthetic effect on postsynaptic glutamatergic neurotransmission. Results:One minimum alveolar concentration sevoflurane depressed the spontaneous activity of 23 inspiratory premotor neurons by (mean ± SD) 30.0 ± 21.0% (P < 0.001). Overall glutamatergic excitation was depressed 19.2 ± 18.5% (P < 0.001), whereas overall GABAAergic inhibition was enhanced by 11.9 ± 25.1% (P < 0.05). The postsynaptic responses to exogenous AMPA and NMDA did not change. Conclusion:One minimum alveolar concentration depressed the activity of inspiratory premotor neurons by a reduction of glutamatergic excitation and an increase in overall inhibition. The postsynaptic AMPA and NMDA receptor response was unchanged. These findings contrast with studies in inspiratory premotor neurons where halothane did not change overall inhibition but significantly reduced the postsynaptic glutamate receptor response.

Effects of Halothane on the Phrenic Nerve Responses to Carbon Dioxide Mediated by Carotid Body Chemoreceptors in Vagotomized Dogs

Background: Previous studies in dogs showed that the phrenic nerve response to an acute hypoxic stimulus was dose dependently depressed by 0.5–2.0 minimum alveolar concentration (MAC) of halothane but not abolished. Because a carbon dioxide stimulus is transduced by a different mechanism in the carotid body chemoreceptors (CBCRs) than is a hypoxic stimulus, inhalational anesthetics may preferentially depress one of these transduction processes, the central neuronal processing, or both, of the integrated responses to these two types of inputs. Methods: Carotid body chemoreceptor stimulation was produced by short (1–1.5 s), bilateral, 100% carbon dioxide in saline infusions into the carotid arteries during neural inspiration in unpremeditated, halothane‐anesthetized, paralyzed, vagotomized dogs during constant mechanical ventilation. The phrenic neurogram quantified the neural inspiratory response. Four protocols were performed in the study: (1) the dose‐dependent effects of halothane anesthesia (0.5–2.0 MAC) during hyperoxic hypercapnia on phrenic nerve activity, (2) the effects of three background levels of the partial pressure of carbon dioxide (PaCO2) on the magnitude of the carbon dioxide infusion responses at 1 MAC halothane, (3) the effects of anesthetic type on the magnitude of the carbon dioxide infusion response, and (4) the effects of CBCR denervation. Results: Peak phrenic nerve activity (PPA) increased significantly during the carbon dioxide‐stimulated phrenic burst in protocols 1–3; after denervation there was no response (protocol 4). Halothane produced a dose‐dependent reduction in the PPA of control and carbon dioxide infusion‐stimulated phrenic bursts and in the net carbon dioxide response. The net PPA responses for the different PaCO2 background levels were not different but were somewhat larger for sodium thiopental anesthesia than for 1.0 MAC halothane. Conclusions: The phrenic nerve response to an acute, severe carbon dioxide stimulus was dose dependently depressed by surgical doses of halothane. The observed responses to carbon dioxide infusion were mediated by the CBCRs because they were eliminated by CBCR denervation. These results suggest that the CBCR transduction and central transmission of the carbon dioxide signal in terms of inspiratory excitatory drive are not abolished at surgical levels of halothane anesthesia.

Halothane Enhances γ-Aminobutyric Acid Receptor Type A Function but Does Not Change Overall Inhibition in Inspiratory Premotor Neurons in a Decerebrate Dog Model

Background Inspiratory premotor neurons in the caudal ventral medulla relay excitatory drive to phrenic and inspiratory intercostal motoneurons in the spinal cord. These neurons are subject to tonic &ggr;-aminobutyric acid type A (GABAA)–mediated (GABAAergic) inhibition. In a previous study, 1 minimum alveolar concentration (MAC) halothane depressed overall glutamatergic excitatory drive but did not change overall inhibitory drive to the neurons. This study investigated in further detail the effects of halothane on GABAAergic inhibition by examining postsynaptic GABAA receptor activity in these neurons. Methods Studies were performed in decerebrate, vagotomized, paralyzed, and mechanically ventilated dogs during hypercapnic hyperoxia. The effect of 1 MAC halothane on extracellularly recorded neuronal activity was measured during localized picoejection of the GABAA receptor antagonist bicuculline and the GABAA agonist muscimol. Complete blockade of GABAergic inhibition by bicuculline allowed estimation of the prevailing overall inhibition of the neuron. The neuronal response to muscimol was used to assess the anesthetic effect on the postsynaptic GABAA receptor function. Results One minimum alveolar concentration halothane depressed the spontaneous activity of 19 inspiratory premotor neurons by 22.9 ± 29.1% (mean ± SD; P < 0.01). Overall excitatory drive was depressed 23.6 ± 16.9% (P < 0.001). Overall GABAergic inhibition was not changed (+8.7 ± 27.5%; P = 0.295), but the postsynaptic GABAA receptor function was increased by 110.3 ± 97.5% (P < 0.001). Conclusion One minimum alveolar concentration halothane greatly enhanced GABAA receptor function on inspiratory premotor neurons but did not change overall synaptic inhibition, indicating that the presynaptic inhibitory input was reduced. Therefore, the anesthetic depression of spontaneous inspiratory premotor neuronal activity in the intact brainstem respiratory network is mainly due to a decrease in overall glutamatergic excitation.