A cranial implant for stabilizing whole-cell patch-clamp recordings in behaving rodents.

BACKGROUND: In vivo patch-clamp recording techniques provide access to the sub- and suprathreshold membrane potential dynamics of individual neurons during behavior. However, maintaining recording stability throughout behavior is a significant challenge, and while methods for head restraint are commonly used to enhance stability, behaviorally related brain movement relative to the skull can severely impact the success rate and duration of whole-cell patch-clamp recordings. NEW METHOD: We developed a low-cost, biocompatible, and 3D-printable cranial implant capable of locally stabilizing brain movement, while permitting equivalent access to the brain when compared to a conventional craniotomy. RESULTS: Experiments in head-restrained behaving mice demonstrate that the cranial implant can reliably reduce the amplitude and speed of brain displacements, significantly improving the success rate of recordings across repeated bouts of motor behavior. COMPARISON WITH EXISTING METHOD(S): Our solution offers an improvement on currently available strategies for brain stabilization. Due to its small size, the implant can be retrofitted to most in vivo electrophysiology recording setups, providing a low cost, easily implementable solution for increasing intracellular recording stability in vivo. CONCLUSIONS: By facilitating stable whole-cell patch-clamp recordings in vivo, biocompatible 3D printed implants should accelerate the investigation of single neuron computations underlying behavior.

Movement-specific signaling is differentially distributed across motor cortex layer 5 projection neuron classes.

Motor cortex generates descending output necessary for executing a wide range of limb movements. Although movement-related activity has been described throughout motor cortex, the spatiotemporal organization of movement-specific signaling in deep layers remains largely unknown. Here we record layer 5B population dynamics in the caudal forelimb area of motor cortex while mice perform a forelimb push/pull task and find that most neurons show movement-invariant responses, with a minority displaying movement specificity. Using cell-type-specific imaging, we identify that invariant responses dominate pyramidal tract (PT) neuron activity, with a small subpopulation representing movement type, whereas a larger proportion of intratelencephalic (IT) neurons display movement-type-specific signaling. The proportion of IT neurons decoding movement-type peaks prior to movement initiation, whereas for PT neurons, this occurs during movement execution. Our data suggest that layer 5B population dynamics largely reflect movement-invariant signaling, with information related to movement-type being routed through relatively small, distributed subpopulations of projection neurons.

A cerebellar-thalamocortical pathway drives behavioral context-dependent movement initiation.

Executing learned motor behaviors often requires the transformation of sensory cues into patterns of motor commands that generate appropriately timed actions. The cerebellum and thalamus are two key areas involved in shaping cortical output and movement, but the contribution of a cerebellar-thalamocortical pathway to voluntary movement initiation remains poorly understood. Here, we investigated how an auditory "go cue" transforms thalamocortical activity patterns and how these changes relate to movement initiation. Population responses in dentate/interpositus-recipient regions of motor thalamus reflect a time-locked increase in activity immediately prior to movement initiation that is temporally uncoupled from the go cue, indicative of a fixed-latency feedforward motor timing signal. Blocking cerebellar or motor thalamic output suppresses movement initiation, while stimulation triggers movements in a behavioral context-dependent manner. Our findings show how cerebellar output, via the thalamus, shapes cortical activity patterns necessary for learned context-dependent movement initiation.

A Temporal Filter for Binaural Hearing Is Dynamically Adjusted by Sound Pressure Level.

In natural environments our auditory system is exposed to multiple and diverse signals of fluctuating amplitudes. Therefore, to detect, localize, and single out individual sounds the auditory system has to process and filter spectral and temporal information from both ears. It is known that the overall sound pressure level affects sensory signal transduction and therefore the temporal response pattern of auditory neurons. We hypothesize that the mammalian binaural system utilizes a dynamic mechanism to adjust the temporal filters in neuronal circuits to different overall sound pressure levels. Previous studies proposed an inhibitory mechanism generated by the reciprocally coupled dorsal nuclei of the lateral lemniscus (DNLL) as a temporal neuronal-network filter that suppresses rapid binaural fluctuations. Here we investigated the consequence of different sound levels on this filter during binaural processing. Our in vivo and in vitro electrophysiology in Mongolian gerbils shows that the integration of ascending excitation and contralateral inhibition defines the temporal properties of this inhibitory filter. The time course of this filter depends on the synaptic drive, which is modulated by the overall sound pressure level and N-methyl-D-aspartate receptor (NMDAR) signaling. In psychophysical experiments we tested the temporal perception of humans and show that detection and localization of two subsequent tones changes with the sound pressure level consistent with our physiological results. Together our data support the hypothesis that mammals dynamically adjust their time window for sound detection and localization within the binaural system in a sound level dependent manner.

Electrogenic N-methyl-D-aspartate receptor signaling enhances binaural responses in the adult brainstem.

In sensory systems, the neuronal representation of external stimuli is enhanced along the sensory pathway. In the auditory system, strong enhancement of binaural information takes place between the brainstem and the midbrain; however, the underlying cellular mechanisms are unknown. Here we investigated the transformation of binaural information in the dorsal nucleus of the lateral lemniscus (DNLL), a nucleus that connects the binaural nuclei in the brainstem and the inferior colliculus in the midbrain. We used in vitro and in vivo electrophysiology in adult Mongolian gerbils to show that N-methyl-D-aspartate receptor (NMDARs) play a critical role in neuronal encoding of stimulus properties in the DNLL. While NMDARs increase firing rates, the timing and the accuracy of the neuronal responses remain unchanged. NMDAR-mediated excitation increases the information about the acoustic stimulus. Taken together, our results show that NMDARs in the DNLL enhance the auditory information content in adult mammal brainstem.

Activity-dependent transmission and integration control the timescales of auditory processing at an inhibitory synapse.

To capture the context of sensory information, neural networks must process input signals across multiple timescales. In the auditory system, a prominent change in temporal processing takes place at an inhibitory GABAergic synapse in the dorsal nucleus of the lateral lemniscus (DNLL). At this synapse, inhibition outlasts the stimulus by tens of milliseconds, such that it suppresses responses to lagging sounds, and is therefore implicated in echo suppression. Here, we untangle the cellular basis of this inhibition. We demonstrate with in vivo whole-cell patch-clamp recordings in Mongolian gerbils that the duration of inhibition increases with sound intensity. Activity-dependent spillover and asynchronous release translate the high presynaptic firing rates found in vivo into a prolonged synaptic output in acute slice recordings. A key mechanism controlling the inhibitory time course is the passive integration of the hyperpolarizing inhibitory conductance. This prolongation depends on the synaptic conductance amplitude. Computational modeling shows that this prolongation is a general mechanism and relies on a non-linear effect caused by synaptic conductance saturation when approaching the GABA reversal potential. The resulting hyperpolarization generates an efficient activity-dependent suppression of action potentials without affecting the threshold or gain of the input-output function. Taken together, the GABAergic inhibition in the DNLL is adjusted to the physiologically relevant duration by passive integration of inhibition with activity-dependent synaptic kinetics. This change in processing timescale combined with the reciprocal connectivity between the DNLLs implements a mechanism to suppress the distracting localization cues of echoes and helps to localize the initial sound source reliably.

Development and modulation of intrinsic membrane properties control the temporal precision of auditory brain stem neurons.

Passive and active membrane properties determine the voltage responses of neurons. Within the auditory brain stem, refinements in these intrinsic properties during late postnatal development usually generate short integration times and precise action-potential generation. This developmentally acquired temporal precision is crucial for auditory signal processing. How the interactions of these intrinsic properties develop in concert to enable auditory neurons to transfer information with high temporal precision has not yet been elucidated in detail. Here, we show how the developmental interaction of intrinsic membrane parameters generates high firing precision. We performed in vitro recordings from neurons of postnatal days 9-28 in the ventral nucleus of the lateral lemniscus of Mongolian gerbils, an auditory brain stem structure that converts excitatory to inhibitory information with high temporal precision. During this developmental period, the input resistance and capacitance decrease, and action potentials acquire faster kinetics and enhanced precision. Depending on the stimulation time course, the input resistance and capacitance contribute differentially to action-potential thresholds. The decrease in input resistance, however, is sufficient to explain the enhanced action-potential precision. Alterations in passive membrane properties also interact with a developmental change in potassium currents to generate the emergence of the mature firing pattern, characteristic of coincidence-detector neurons. Cholinergic receptor-mediated depolarizations further modulate this intrinsic excitability profile by eliciting changes in the threshold and firing pattern, irrespective of the developmental stage. Thus our findings reveal how intrinsic membrane properties interact developmentally to promote temporally precise information processing.

Large somatic synapses on neurons in the ventral lateral lemniscus work in pairs.

In the auditory system, large somatic synapses convey strong excitation that supports temporally precise information transfer. The information transfer of such synapses has predominantly been investigated in the endbulbs of Held in the anterior ventral cochlear nucleus and the calyx of Held in the medial nucleus of the trapezoid body. These large synapses either work as relays or integrate over a small number of inputs to excite the postsynaptic neuron beyond action potential (AP) threshold. In the monaural system, another large somatic synapse targets neurons in the ventral nucleus of the lateral lemniscus (VNLL). Here, we comparatively analyze the mechanisms of synaptic information transfer in endbulbs in the VNLL and the calyx of Held in juvenile Mongolian gerbils. We find that endbulbs in the VNLL are functionally surface-scaled versions of the calyx of Held with respect to vesicle availability, release efficacy, and synaptic peak currents. This functional scaling is achieved by different calcium current kinetics that compensate for the smaller AP in VNLL endbulbs. However, the average postsynaptic current in the VNLL fails to elicit APs in its target neurons, even though equal current suffices to generate APs in neurons postsynaptic to the calyx of Held. In the VNLL, a postsynaptic A-type outward current reduces excitability and prevents AP generation upon a single presynaptic input. Instead, coincidence detection of inputs from two converging endbulbs is ideal to reliably trigger APs. Thus, even large endbulbs do not guarantee one-to-one AP transfer. Instead, information flow appears regulated by circuit requirements.