Bryan M. Krause

Preferential effect of isoflurane on top-down vs. bottom-up pathways in sensory cortex

The mechanism of loss of consciousness (LOC) under anesthesia is unknown. Because consciousness depends on activity in the cortico-thalamic network, anesthetic actions on this network are likely critical for LOC. Competing theories stress the importance of anesthetic actions on bottom-up “core” thalamo-cortical (TC) vs. top-down cortico-cortical (CC) and matrix TC connections. We tested these models using laminar recordings in rat auditory cortex in vivo and murine brain slices. We selectively activated bottom-up vs. top-down afferent pathways using sensory stimuli in vivo and electrical stimulation in brain slices, and compared effects of isoflurane on responses evoked via the two pathways. Auditory stimuli in vivo and core TC afferent stimulation in brain slices evoked short latency current sinks in middle layers, consistent with activation of core TC afferents. By contrast, visual stimuli in vivo and stimulation of CC and matrix TC afferents in brain slices evoked responses mainly in superficial and deep layers, consistent with projection patterns of top-down afferents that carry visual information to auditory cortex. Responses to auditory stimuli in vivo and core TC afferents in brain slices were significantly less affected by isoflurane compared to responses triggered by visual stimuli in vivo and CC/matrix TC afferents in slices. At a just-hypnotic dose in vivo, auditory responses were enhanced by isoflurane, whereas visual responses were dramatically reduced. At a comparable concentration in slices, isoflurane suppressed both core TC and CC/matrix TC responses, but the effect on the latter responses was far greater than on core TC responses, indicating that at least part of the differential effects observed in vivo were due to local actions of isoflurane in auditory cortex. These data support a model in which disruption of top-down connectivity contributes to anesthesia-induced LOC, and have implications for understanding the neural basis of consciousness.

Descending Projections from Extrastriate Visual Cortex Modulate Responses of Cells in Primary Auditory Cortex

Primary sensory cortical responses are modulated by the presence or expectation of related sensory information in other modalities, but the sources of multimodal information and the cellular locus of this integration are unclear. We investigated the modulation of neural responses in the murine primary auditory cortical area Au1 by extrastriate visual cortex (V2). Projections from V2 to Au1 terminated in a classical descending/modulatory pattern, with highest density in layers 1, 2, 5, and 6. In brain slices, whole-cell recordings revealed long latency responses to stimulation in V2L that could modulate responses to subsequent white matter (WM) stimuli at latencies of 5-20 ms. Calcium responses imaged in Au1 cell populations showed that preceding WM with V2L stimulation modulated WM responses, with both summation and suppression observed. Modulation of WM responses was most evident for near-threshold WM stimuli. These data indicate that corticocortical projections from V2 contribute to multimodal integration in primary auditory cortex.

Spiking in auditory cortex following thalamic stimulation is dominated by cortical network activity.

The state of the sensory cortical network can have a profound impact on neural responses and perception. In rodent auditory cortex, sensory responses are reported to occur in the context of network events, similar to brief UP states, that produce “packets” of spikes and are associated with synchronized synaptic input (Bathellier et al., 2012; Hromadka et al., 2013; Luczak et al., 2013). However, traditional models based on data from visual and somatosensory cortex predict that ascending sensory thalamocortical (TC) pathways sequentially activate cells in layers 4 (L4), L2/3, and L5. The relationship between these two spatio-temporal activity patterns is unclear. Here, we used calcium imaging and electrophysiological recordings in murine auditory TC brain slices to investigate the laminar response pattern to stimulation of TC afferents. We show that although monosynaptically driven spiking in response to TC afferents occurs, the vast majority of spikes fired following TC stimulation occurs during brief UP states and outside the context of the L4>L2/3>L5 activation sequence. Specifically, monosynaptic subthreshold TC responses with similar latencies were observed throughout layers 2–6, presumably via synapses onto dendritic processes located in L3 and L4. However, monosynaptic spiking was rare, and occurred primarily in L4 and L5 non-pyramidal cells. By contrast, during brief, TC-induced UP states, spiking was dense and occurred primarily in pyramidal cells. These network events always involved infragranular layers, whereas involvement of supragranular layers was variable. During UP states, spike latencies were comparable between infragranular and supragranular cells. These data are consistent with a model in which activation of auditory cortex, especially supragranular layers, depends on internally generated network events that represent a non-linear amplification process, are initiated by infragranular cells and tightly regulated by feed-forward inhibitory cells.

Disruption of cortical network activity by the general anaesthetic isoflurane

Background Actions of general anaesthetics on activity in the cortico-thalamic network likely contribute to loss of consciousness and disconnection from the environment. Previously, we showed that the general anaesthetic isoflurane preferentially suppresses cortically evoked synaptic responses compared with thalamically evoked synaptic responses, but how this differential sensitivity translates into changes in network activity is unclear. Methods We investigated isoflurane disruption of spontaneous and stimulus-induced cortical network activity using multichannel recordings in murine auditory thalamo-cortical brain slices. Results Under control conditions, afferent stimulation elicited short latency, presumably monosynaptically driven, spiking responses, as well as long latency network bursts that propagated horizontally through the cortex. Isoflurane (0.05-0.6 mM) suppressed spiking activity overall, but had a far greater effect on network bursts than on early spiking responses. At isoflurane concentrations >0.3 mM, network bursts were almost entirely blocked, even with increased stimulation intensity and in response to paired (thalamo-cortical + cortical layer 1) stimulation, while early spiking responses were <50% blocked. Isoflurane increased the threshold for eliciting bursts, decreased their propagation speed and prevented layer 1 afferents from facilitating burst induction by thalamo-cortical afferents. Conclusions Disruption of horizontal activity spread and of layer 1 facilitation of thalamo-cortical responses likely contribute to the mechanism by which suppression of cortical feedback connections disrupts sensory awareness under anaesthesia.

Respiratory neuron characterization reveals intrinsic bursting properties in isolated adult turtle brainstems (Trachemys scripta).

It is not known whether respiratory neurons with intrinsic bursting properties exist within ectothermic vertebrate respiratory control systems. Thus, isolated adult turtle brainstems spontaneously producing respiratory motor output were used to identify and classify respiratory neurons based on their firing pattern relative to hypoglossal (XII) nerve activity. Most respiratory neurons (183/212) had peak activity during the expiratory phase, while inspiratory, post-inspiratory, and novel pre-expiratory neurons were less common. During synaptic blockade conditions, ∼10% of respiratory neurons fired bursts of action potentials, with post-inspiratory cells (6/9) having the highest percentage of intrinsic burst properties. Most intrinsically bursting respiratory neurons were clustered at the level of the vagus (X) nerve root. Synaptic inhibition blockade caused seizure-like activity throughout the turtle brainstem, which shows that the turtle respiratory control system is not transformed into a network driven by intrinsically bursting respiratory neurons. We hypothesize that intrinsically bursting respiratory neurons are evolutionarily conserved and represent a potential rhythmogenic mechanism contributing to respiration in adult turtles.

Altered stimulus representation in rat auditory cortex is not causal for loss of consciousness under general anaesthesia

Background: Current concepts suggest that impaired representation of information in cortical networks contributes to loss of consciousness under anaesthesia. We tested this idea in rat auditory cortex using information theory analysis of multiunit responses recorded under three anaesthetic agents with different molecular targets: isoflurane, propofol, and dexmedetomidine. We reasoned that if changes in the representation of sensory stimuli are causal for loss of consciousness, they should occur regardless of the specific anaesthetic agent. Methods: Spiking responses were recorded with chronically implanted microwire arrays in response to acoustic stimuli incorporating varied temporal and spectral dynamics. Experiments consisted of four drug conditions: awake (pre‐drug), sedation (i.e. intact righting reflex), loss of consciousness (a dose just sufficient to cause loss of righting reflex), and recovery. Measures of firing rate, spike timing, and mutual information were analysed as a function of drug condition. Results: All three drugs decreased spontaneous and evoked spiking activity and modulated spike timing. However, changes in mutual information were inconsistent with altered stimulus representation being causal for loss of consciousness. First, direction of change in mutual information was agent‐specific, increasing under dexmedetomidine and decreasing under isoflurane and propofol. Second, mutual information did not decrease at the transition between sedation and LOC for any agent. Changes in mutual information under anaesthesia correlated strongly with changes in precision and reliability of spike timing, consistent with the importance of temporal stimulus features in driving auditory cortical activity. Conclusions: The primary sensory cortex is not the locus for changes in representation of information causal for loss of consciousness under anaesthesia.

PV+ Cells Enhance Temporal Population Codes but not Stimulus-Related Timing in Auditory Cortex

Spatio-temporal cortical activity patterns relative to both peripheral input and local network activity carry information about stimulus identity and context. GABAergic interneurons are reported to regulate spiking at millisecond precision in response to sensory stimulation and during gamma oscillations; their role in regulating spike timing during induced network bursts is unclear. We investigated this issue in murine auditory thalamo-cortical (TC) brain slices, in which TC afferents induced network bursts similar to previous reports in vivo. Spike timing relative to TC afferent stimulation during bursts was poor in pyramidal cells and SOM+ interneurons. It was more precise in PV+ interneurons, consistent with their reported contribution to spiking precision in pyramidal cells. Optogenetic suppression of PV+ cells unexpectedly improved afferent-locked spike timing in pyramidal cells. In contrast, our evidence suggests that PV+ cells do regulate the spatio-temporal spike pattern of pyramidal cells during network bursts, whose organization is suited to ensemble coding of stimulus information. Simulations showed that suppressing PV+ cells reduces the capacity of pyramidal cell networks to produce discriminable spike patterns. By dissociating temporal precision with respect to a stimulus versus internal cortical activity, we identified a novel role for GABAergic cells in regulating information processing in cortical networks.

Analysis of stimulus-related activity in rat auditory cortex using complex spectral coefficients.

The neural mechanisms of sensory responses recorded from the scalp or cortical surface remain controversial. Evoked vs. induced response components (i.e., changes in mean vs. variance) are associated with bottom-up vs. top-down processing, but trial-by-trial response variability can confound this interpretation. Phase reset of ongoing oscillations has also been postulated to contribute to sensory responses. In this article, we present evidence that responses under passive listening conditions are dominated by variable evoked response components. We measured the mean, variance, and phase of complex time-frequency coefficients of epidurally recorded responses to acoustic stimuli in rats. During the stimulus, changes in mean, variance, and phase tended to co-occur. After the stimulus, there was a small, low-frequency offset response in the mean and modest, prolonged desynchronization in the alpha band. Simulations showed that trial-by-trial variability in the mean can account for most of the variance and phase changes observed during the stimulus. This variability was state dependent, with smallest variability during periods of greatest arousal. Our data suggest that cortical responses to auditory stimuli reflect variable inputs to the cortical network. These analyses suggest that caution should be exercised when interpreting variance and phase changes in terms of top-down cortical processing.

Evidence that loss of consciousness under anesthesia is not associated with impaired stimulus representation in auditory cortex

The mechanism whereby anesthetics cause loss of consciousness (LOC) is poorly understood. Current theories suggest that impaired representation of information in cortico-thalamic networks contributes to LOC under anesthesia. We sought to determine whether such changes are present in auditory cortex using information theoretic analysis of multiunit responses in rats. We tested the effects of three agents with different molecular targets: isoflurane, which acts at multiple pre- and postsynaptic loci, propofol, which acts primarily on GABAA receptors, and dexmedetomidine, an α2 adrenergic agonist. We reasoned that changes in the representation of sensory stimuli causative for LOC would be present regardless of the molecular target of the anesthetic. All three agents caused LOC, as assayed by the loss of righting reflex (LORR). We presented acoustic stimuli that varied across a wide range of temporal and spectral dynamics under control, sub-hypnotic (i.e. dose too low to cause LORR), just-hypnotic (a dose just sufficient to cause LORR) and recovery conditions. Changes in mutual information (MI) between the stimulus and spike responses under anesthesia diverged in two ways from predictions of a model in which stimulus representation is impaired upon LOC. First, the sign of changes in MI was agent-specific: MI increased under dexmedetomidine, while it decreased under isoflurane and propofol. Second, there was no consistent change in MI when transitioning from sub-hyptnotic to just-hypnotic doses: for none of the agents did MI decrease at the higher dose, and in some cases MI actually increased relative to the sub-hypnotic dose. Changes in MI under anesthesia were strongly correlated with changes in precision and reliability of spike timing, consistent with the importance of temporal stimulus features in driving auditory cortical activity. These data indicate that primary sensory cortex is not the locus for changes in information representation causative for LOC under anesthesia.

Evidence that PV+ cells enhance temporal population codes but not stimulus-related timing in auditory cortex

Spatio-temporal cortical activity patterns relative to both peripheral input and local network activity carry information about stimulus identity and context. GABAergic interneurons are reported to regulate spiking at millisecond precision in response to sensory stimulation and during gamma oscillations; their role in regulating spike timing during induced network bursts is unclear. We investigated this issue in murine auditory thalamo-cortical (TC) brain slices, in which TC afferents induced network bursts similar to previous reports in vivo. Spike timing relative to TC afferent stimulation during bursts was poor in pyramidal cells and SOM+ interneurons. It was more precise in PV+ interneurons, consistent with their reported contribution to spiking precision in pyramidal cells. Optogenetic suppression of PV+ cells unexpectedly improved afferent-locked spike timing in pyramidal cells. In contrast, our evidence suggests that PV+ cells do regulate the spatio-temporal spike pattern of pyramidal cells during network bursts, whose organization is suited to ensemble coding of stimulus information. Simulations showed that suppressing PV+ cells reduces the capacity of pyramidal cell networks to produce discriminable spike patterns. By dissociating temporal precision with respect to a stimulus versus internal cortical activity, we identified a novel role for GABAergic cells in regulating information processing in cortical networks.