Mislav Tonkovic-Capin

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 Sevoflurane on excitatory neurotransmission to medullary expiratory neurons and on phrenic nerve activity in a decerebrate dog model

Background Sevoflurane is a new volatile anesthetic with a pronounced respiratory depressant effect. Synaptic neurotransmission in canine expiratory bulbospinal neurons is mainly mediated by excitatory N-methyl-d-aspartatic acid (NMDA) receptor input and modulated by inhibitory &ggr;-aminobutyric acid type A (GABAA) receptors. The authors investigated the effect of sevoflurane on these 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 (MAC; 2.4%) sevoflurane on extracellularly recorded neuronal activity was measured during localized picoejection of the glutamate agonist NMDA and the GABAA receptor blocker bicuculline in a two-part protocol. First, complete blockade of the GABAAergic mechanism by bicuculline allowed differentiation between the effects of sevoflurane on overall GABAAergic inhibition and on overall glutamatergic excitation. In a second step, the neuronal response to exogenous NMDA was used to estimate sevoflurane’s effect on postsynaptic glutamatergic neurotransmission. Results One minimum alveolar concentration sevoflurane depressed the spontaneous activity of 16 expiratory neurons by 36.7 ± 22.4% (mean ± SD). Overall glutamatergic excitation was depressed 19.5 ± 16.2%, and GABAAergic inhibition was enhanced 18.7 ± 20.6%. However, the postsynaptic response to exogenous NMDA was not significantly altered. In addition, 1 MAC sevoflurane depressed peak phrenic nerve activity by 61.8 ± 17.7%. Conclusions In the authors’in vivo expiratory neuronal model, the depressive effect of sevoflurane on synaptic neurotransmission was caused by a reduction of presynaptic glutamatergic excitation and an enhancement of GABAAergic inhibition. The effects on expiratory neuronal activity were similar to halothane, but sevoflurane caused a stronger depression of phrenic nerve activity than halothane.

Dose-dependent Effects of Halothane on the Carbon Dioxide Responses of Expiratory and Inspiratory Bulbospinal Neurons and the Phrenic Nerve Activities in Dogs

Background:Expiratory bulbospinal and inspiratory bulbospinal neurons in the ventral respiratory group provide drive for thoracoabdominal expiratory and phrenic and thoracic inspiratory motor neurons. Potent inhalational agents such as halothane may have differential effects on inspiratory and expiratory neurons, but detailed studies comparing neurons at a homologous level are lacking. Methods:The dose-dependent effects of anesthesia with 1.0- 2.5 minimum alveolar concentration halothane on the CO2 responses of single expiratory and inspiratory bulbospinal neurons of the ventral respiratory group and on phrenic neural activities were studied in nonpremedicated, anesthetized, paralyzed, vagotomlzed dogs. Hyperventllation with O2 and the addition of CO2-O2 mixtures were used to produce low, medium, and high steady-state levels of central CO2 drive. Results:Peak neuron discharge frequency decreased progressively with increasing halothane dose at all levels of CO2 drive for both types of neurons. The sensitivities of inspiratory and expiratory bulbospinal neuronal activities to halothane were not significantly different from one another, whereas the sensitivity to halothane of the peak phrenic activity was markedly greater than those of the neurons. Increasing halothane dose caused a downward, predominantly parallel shift of the CO2 response curves. Phrenic nerve activity also showed a decrease in slope of the CO2 response. Conclusions:The activities of respiratory premotor neurons are less depressed by Increasing doses of halothane than is phrenic nerve activity. The greater depression of phrenic activity may result from additional anesthetic actions on the efferent motor pathways, resulting in decreased descending synaptic inputs to phrenic motor neurons.

Dose-dependent effects of isoflurane on the CO2 responses of expiratory medullary neurons and the phrenic nerve activities in dogs.

Expiratory bulbospinal neurons (EBS) neurons in the region of the nucleus retroambigualis provide a major source of drive for thoracic and abdominal expiratory motoneurons. These studies examined the dose-dependent effects of isoflurane anesthesia from 0.5-2.5 minimum alveolar concentration (MAC) on the CO2 responses of extracellularly recorded single EBS neurons and phrenic neural activities in unpremedicated, anesthetized, paralyzed, vagotomized dogs. Hyperventilation with O2 and the addition of CO2-O2 mixtures were used to produce three steady-state levels of central CO2 drive: low, medium, and high with corresponding mean PaCO2 values of 29, 45, and 67 mmHg. Plots of peak expiratory neuron discharge frequency (Fn) versus steady-state levels of isoflurane dose showed a progressive decrease in Fn for both high and medium drive levels, whereas for low drive most neurons fired tonically at all MAC levels with little change in Fn. Peak phrenic activity showed a similar but more pronounced decline with increased MAC for all levels of CO2 drive. Increasing isoflurane dose caused a downward, predominantly parallel shift of the CO2 response curves for both expiratory neuronal and phrenic neural activities. Phrenic activity ceased at 2-2.5 MAC isoflurane for all CO2 drive levels, whereas EBS neurons continued to fire tonically at the same depth at 30-50% of Fn at 1 MAC for the high level. These studies suggest that phrenic (inspiratory) activity is more sensitive than EBS neurons to isoflurane and that increases in CO2 drive can only partially offset the depressant effect of isoflurane on central respiratory activities.

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.

Opioid Receptors on Bulbospinal Respiratory Neurons Are Not Activated During Neuronal Depression by Clinically Relevant Opioid Concentrations

Opioids depress the activity of brain stem respiratory-related neurons, but it is not resolved whether the mechanism at clinical concentrations consists of direct neuronal effects or network effects. We performed extracellular recordings of discharge activity of single respiratory neurons in the caudal ventral respiratory group of decerebrate dogs, which were premotor neurons with a likelihood of 90%. We used multibarrel glass microelectrodes, which allowed concomitant highly localized picoejection of opioid receptor agonists or antagonists onto the neuron. Picoejection of the mu receptor agonist [d-Ala(2), N-Me-phe(4), gly-ol(5)]-enkephalin (DAMGO, 1 mM) decreased the peak discharge frequency (mean +/- SD) of expiratory neurons to 68 +/- 22% (n = 12), the delta(1) agonist d-Pen(2,5)-enkephalin (DPDPE, 1 mM) to 95 +/- 11% (n = 15), and delta(2) receptor agonist [d-Ala(2)] deltorphin-II to 86 +/- 17% (1 mM, n = 15). The corresponding values for inspiratory neurons were: 64 +/- 12% (n = 11), 48 +/- 30% (n = 12), and 75 +/- 15% (n = 11), respectively. Naloxone fully reversed these effects. Picoejection of morphine (0.01-1 mM) depressed most neurons in a concentration dependent fashion to maximally 63% (n = 27). Picoejection of remifentanil (240-480 nM) did not cause any significant depression of inspiratory (n = 11) or expiratory neurons (n = 9). 4. Intravenous remifentanil (0.2-0.6 microg.kg(-1).min(-1)) decreased neuronal peak discharge frequency to 60 +/- 12% (inspiratory, n = 7) and 58 +/- 11% (expiratory, n = 11). However, local picoejection of naloxone did not reverse the neuronal depression. Our data suggest that mu, delta(1), and delta(2) receptors are present on canine respiratory premotor neurons. Clinical concentrations of morphine and remifentanil caused no local depression. This lack of effect and the inability of local naloxone to reverse the neuronal depression by intravenous remifentanil suggest that clinical concentrations of opioids produce their depressive effects on mechanisms upstream from respiratory bulbospinal premotor neurons.

Effects of halothane and sevoflurane on inhibitory neurotransmission to medullary expiratory neurons in a decerebrate dog model.

Background In canine expiratory bulbospinal neurons, 1 minimum alveolar concentration (MAC) halothane and sevoflurane reduced the glutamatergic excitatory drive at a presynaptic site and enhanced the overall &ggr;-aminobutyric acid (GABA)-mediated inhibitory input. The authors investigated if this inhibitory enhancement was mainly caused by postsynaptic effects. Methods Two separate anesthetic studies were performed in two sets of decerebrate, vagotomized, paralyzed, and mechanically ventilated dogs during hypercapnic hyperoxia. The effect of 1 MAC halothane or sevoflurane on extracellularly recorded neuronal activity was measured during localized picoejection of the GABAA receptor agonist muscimol and the GABAA receptor antagonist bicuculline. Complete blockade of GABAA-mediated inhibition with bicuculline was used to assess the prevailing overall inhibitory input to the neuron. The neuronal response to muscimol was used to estimate the anesthetic effect on postsynaptic GABAA receptor function. Results Halothane at 1 MAC depressed the spontaneous activity of 12 expiratory neurons 22.2 ± 14.8% (mean ± SD) and overall glutamatergic excitation 14.5 ± 17.9%. Overall GABA-mediated inhibition was enhanced 14.1 ± 17.9% and postsynaptic GABAA receptor function 74.2 ± 69.2%. Sevoflurane at 1 MAC depressed the spontaneous activity of 23 neurons 20.6 ± 19.3% and overall excitation 10.6 ± 21.7%. Overall inhibition was enhanced 15.4 ± 34.0% and postsynaptic GABAA receptor function 65.0 ± 70.9%. The effects of halothane and sevoflurane were not statistically different. Conclusion Halothane and sevoflurane at 1 MAC produced a small increase in overall inhibition of expiratory premotor neuronal activity. The increase in inhibition results from a marked enhancement of postsynaptic GABAA receptor function that is partially offset by a reduction in presynaptic inhibitory input by the anesthetics.

Halothane Depresses Glutamatergic Neurotransmission to Brain Stem Inspiratory Premotor Neurons in a Decerebrate Dog Model

Background Inspiratory bulbospinal neurons in the caudal ventral medulla are premotor neurons that drive phrenic motoneurons and ultimately the diaphragm. 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 modulated by an inhibitory &ggr;-aminobutyric acidA (GABAA)ergic input. The authors investigated the effect of halothane 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 (MAC) halothane on extracellularly recorded neuronal activity 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 halothane 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 Halothane, 1 MAC, depressed the spontaneous activity of 21 inspiratory neurons by 20.6 ± 18.0% (mean ± SD;P = 0.012). Overall glutamatergic excitation was depressed 15.4 ± 20.2% (P = 0.001), while overall GABAAergic inhibition did not change. The postsynaptic responses to exogenous AMPA and NMDA were also depressed by 18.6 ± 35.7% (P = 0.03) and 22.2 ± 26.2% (P = 0.004), respectively. Conclusion Halothane, 1 MAC, depressed the activity of inspiratory premotor neurons by a reduction of glutamatergic excitation. Overall inhibitory drive did not change. The postsynaptic AMPA and NMDA receptor response was significantly reduced. These findings contrast with studies in expiratory premotor neurons in which overall inhibition was significantly increased by halothane and there was no reduction in the postsynaptic glutamate receptor response.