Tonic Firing Research Articles

Spiking patterns and synchronization of thalamic neurons along the sleep-wake cycle

Spiking patterns and synchronization dynamics of thalamic neurons along the sleep-wake cycle are studied in a minimal model of four coupled conductance-based neurons. The model simulates two thalamic neurons coupled via a gap junction and driven by a synaptic input from a two-neuron model of sleep regulation by the hypothalamus. In accord with experimental data, the model shows that during sleep, when hypothalamic wake-active neurons are silent, the thalamic neurons discharge bursts of spikes. During wake, the excitatory synaptic input from the hypothalamus drives the coupled thalamic neurons to a state of tonic firing (single spikes). In the deterministic case, the thalamic neurons synchronize in-phase in the bursting regime but demonstrate multi-stability of out-of-phase, in-phase, and asynchronous states in the tonic firing. However, along the sleep-wake cycle, once the neurons synchronize in-phase during sleep (bursting), they stay synchronized in wake (tonic firing). It is thus found that noise is needed to reproduce the experimentally observed transitions between synchronized bursting during sleep and asynchronous tonic firing during wake. Overall, synchronization of bursting is found to be more robust to noise than synchronization of tonic firing, where a small disturbance is sufficient to desynchronize the thalamic neurons. The model predicts that the transitions between sleep and wake happen via chaos because a single thalamic neuron exhibits chaos between regular bursting and tonic activity. The results of this study suggest that the sleep- and wake-related dynamics in the thalamus may be generated at a level of gap junction-coupled clusters of thalamic neurons driven from the hypothalamus which would then propagate throughout the thalamus and cortex via axonal long-range connections.

Activity patterns with silent states in a heterogeneous network of gap-junction coupled Huber-Braun model neurons.

A mathematical model of a network of nearest neighbor gap-junction coupled neurons has been used to examine the impact of neuronal heterogeneity on the networks' activity during increasing coupling strength. Heterogeneity has been introduced by Huber-Braun model neurons with randomization of the temperature as a scaling factor. This leads to neurons of an enormous diversity of impulse pattern, including burst discharges, chaotic activity, and two different types of tonic firing-all of them experimentally observed in the peripheral as well as central nervous system. When the network is composed of all these types of neurons, randomly selected, a particular phenomenon can be observed. At a certain coupling strength, the network goes into a completely silent state. Examination of voltage traces and inter-spike intervals of individual neurons suggests that all neurons, irrespective of their original pattern, go through a well-known bifurcation scenario, resembling those of single neurons especially on external current injection. All the originally spontaneously firing neurons can achieve constant membrane potentials at which all intrinsic and gap-junction currents are balanced. With limited diversity, i.e., taking out neurons of specific patterns from the lower and upper temperature range, spontaneous firing can be reinstalled with further increasing coupling strength, especially when the tonic firing regimes are missing. Reinstalled firing develops from slowly increasing subthreshold oscillations leading to tonic firing activity with already fairly well synchronized action potentials, while the subthreshold potentials can still be significantly different. Full in phase synchronization is not achieved. Additional studies are needed elucidating the underlying mechanisms and the conditions under which such particular transitions can appear.

Gabapentin regulates dopaminergic neuron firing and theta oscillation in the ventral tegmental area to reverse depression-like behavior in chronic neuropathic pain state.

PurposeVentral tegmental area (VTA) dopamine system plays an important role in depression and is also involved in pain experience. In this study, we investigated the VTA dopaminergic (DA) neuron firing and local field potential (LFP) in pain-related depression, and we try to explore the underlying relationship between pain and depression.Materials and methodsWe used neuropathic pain model [spare nerve injury (SNI)] to induce pain-related depression. The Dixon up–down method was used to test mechanical hypersensitivity. Behavioral changes like open field test, sucrose preference test, and forced swim test were used to test depression-like behaviors. Gabapentin (GBP) was used to explore the chronic analgesic treatment that could reverse pain-related depression. To investigate the in vivo variations of VTA DA neuron firing and LFP, multichannel acquisition processor system was used.ResultsWe used SNI to induce depression-like behaviors. Repeated GBP treatment reversed these behaviors after 14 days of injection. An in vivo electrophysiological analysis of the firing characteristics of VTA DA neurons and LFP revealed that SNI increased the firing rate of DA neurons, but not the burst firing activity. Surprisingly, chronic GBP reversed the firing rate of DA neurons and reduced the burst firing activity. Moreover, SNI increased the LFP power in delta and theta oscillation and decreased it in beta oscillation. Repeated administration of GBP significantly suppressed theta oscillation. Above all, chronic GBP altered these characteristics to reverse depression-like behaviors.ConclusionThe present study confirmed that the tonic firing activity of VTA DA neurons, but not the burst firing activity, was the key factor in peripheral neuropathy–induced depression. Chronic GBP regulated the firing pattern of DA neurons and decreased theta oscillation in VTA to treat pain-related depression. This variation tendency of electrophysiological characteristics of VTA DA neurons and theta oscillation in VTA might represent an attempt to cope with pain-related negative mood disorder.

Acute dopamine receptor blockade in substantia nigra pars reticulata: a possible model for drug-induced Parkinsonism.

Dopamine (DA) depletion modifies the firing pattern of neurons in the substantia nigra pars reticulata (SNr), shifting their mostly tonic firing toward irregularity and bursting, traits of pathological firing underlying rigidity and postural instability in Parkinson's disease (PD) patients and animal models of Parkinsonism (PS). Drug-induced Parkinsonism (DIP) represents 20-40% of clinical cases of PS, becoming a problem for differential diagnosis, and is still not well studied with physiological tools. It may co-occur with tardive dyskinesia. Here we use in vitro slice preparations including the SNr to observe drug-induced pathological firing by using drugs that most likely produce it, DA-receptor antagonists (SCH23390 plus sulpiride), to compare with firing patterns found in DA-depleted tissue. The hypothesis is that SNr firing would be similar under both conditions, a prerequisite to the proposal of a similar preparation to test other DIP-producing drugs. Firing was analyzed with three complementary metrics, showing similarities between DA depletion and acute DA-receptor blockade. Moreover, blockade of either nonselective cationic channels or Cav3 T-type calcium channels hyperpolarized the membrane and abolished bursting and irregular firing, silencing SNr neurons in both conditions. Therefore, currents generating firing in control conditions are in part responsible for pathological firing. Haloperidol, a DIP-producing drug, reproduced DA-receptor antagonist firing modifications. Since acute DA-receptor blockade induces SNr neuron firing similar to that found in the 6-hydroxydopamine model of PS, output basal ganglia neurons may play a role in generating DIP. Therefore, this study opens the way to test other DIP-producing drugs. NEW & NOTEWORTHY Dopamine (DA) depletion enhances substantia nigra pars reticulata (SNr) neuron bursting and irregular firing, hallmarks of Parkinsonism. Several drugs, including antipsychotics, antidepressants, and calcium channel antagonists, among others, produce drug-induced Parkinsonism. Here we show the first comparison between SNr neuron firing after DA depletion vs. firing found after acute blockade of DA receptors. It was found that firing in both conditions is similar, implying that pathological SNr neuron firing is also a physiological correlate of drug-induced Parkinsonism.

Prefrontal cortex modulates firing pattern in the nucleus reuniens of the midline thalamus via distinct corticothalamic pathways.

The thalamus has long been recognized for its role in relaying sensory information from the periphery, a function accomplished by its "first-order" nuclei. However, a second category of thalamic nuclei, termed "higher-order" nuclei, have been shown instead to mediate communication between cortical areas. The nucleus reuniens of the midline thalamus (RE) is a higher-order nucleus known to act as a conduit of reciprocal communication between the medial prefrontal cortex (mPFC) and hippocampus. While anatomical and behavioural studies of RE are numerous, circuit-based electrophysiological studies, particularly those examining the impact of cortical input and the thalamic reticular nucleus (TRN) on RE neuron firing, are sparse. To characterize RE neuron firing properties and dissect the circuit dynamics of the infralimbic subdivision of the mPFC (ilPFC), the TRN and RE, we used in vivo, extracellular, single-unit recordings in male Sprague Dawley rats and manipulated neural activity using targeted pharmacological manipulations, electrical stimulation and a projection-specific implementation of designer receptors exclusively activated by designer drugs (DREADDs). We show that ilPFC inhibition reduces multiple burst firing parameters in RE, whereas ilPFC stimulation drives burst firing and dampens tonic firing. In addition, TRN inhibition reduces the number of spontaneously active neurons in RE. Finally, inhibition of ilPFC terminals in RE selectively enhances a subset of burst firing parameters. These findings demonstrate that ilPFC input, both via direct projections and via the TRN, can modulate RE neuron firing pattern in nuanced and complex ways. They also highlight the ilPFC-TRN-RE circuit as a likely critical component of prefrontal-hippocampal interactions.

Ionic mechanisms underlying tonic and burst firing behavior in subfornical organ neurons: a combined experimental and modeling study.

Subfornical organ (SFO) neurons exhibit heterogeneity in current expression and spiking behavior, where the two major spiking phenotypes appear as tonic and burst firing. Insight into the mechanisms behind this heterogeneity is critical for understanding how the SFO, a sensory circumventricular organ, integrates and selectively influences physiological function. To integrate efficient methods for studying this heterogeneity, we built a single-compartment, Hodgkin-Huxley-type model of an SFO neuron that is parameterized by SFO-specific in vitro patch-clamp data. The model accounts for the membrane potential distribution and spike train variability of both tonic and burst firing SFO neurons. Analysis of model dynamics confirms that a persistent Na+ and Ca2+ currents are required for burst initiation and maintenance and suggests that a slow-activating K+ current may be responsible for burst termination in SFO neurons. Additionally, the model suggests that heterogeneity in current expression and subsequent influence on spike afterpotential underlie the behavioral differences between tonic and burst firing SFO neurons. Future use of this model in coordination with single neuron patch-clamp electrophysiology provides a platform for explaining and predicting the response of SFO neurons to various combinations of circulating signals, thus elucidating the mechanisms underlying physiological signal integration within the SFO. NEW & NOTEWORTHY Our understanding of how the subfornical organ (SFO) selectively influences autonomic nervous system function remains incomplete but theoretically results from the electrical responses of SFO neurons to physiologically important signals. We have built a computational model of SFO neurons, derived from and supported by experimental data, which explains how SFO neurons produce different electrical patterns. The model provides an efficient system to theoretically and experimentally explore how changes in the essential features of SFO neurons affect their electrical activity.

Effect of 17β-oestradiol on T-type calcium channels in the lateral habenula.

T-type calcium channels (T-channels) are critical for regulating neuronal excitability. Oestrogen alters neuronal excitability by modulating the expression of T-channels. The lateral habenula (LHb), as a link between the limbic system and midbrain structures, expresses T-channels and ERs. However, little is known about the role of oestrogen with respect to modulating T-channels in the LHb. In the present study, we investigated the distribution of T-channels in 3 LHb subregions (rostral, middle and caudal) in normal female rats. Next, we analysed the influence of 17β-oestradiol (E2 ) on T-channels in the LHb in ovariectomised (OVX) rats (oil and E2 groups) using whole-cell patch clamp recording and a real-time polymerase chain reaction (PCR). In normal rats, the results obtained showed that the peak of T-type calcium current (IT ) was -474.61±48.33pA and IT density was -29.11±1.93pA/pF. The IT peak and IT density on LHb neurones gradually decreased across the rostrocaudal axis. The neuronal firing pattern varied depending on the location: burst firing was dominant (53.85%) in the rostral LHb, whereas tonic firing was dominant (79.31%) in the caudal LHb. In OVX rats, real-time PCR analysis revealed that E2 treatment decreased Cav3.3 mRNA expression in the caudal LHb. Patch clamp recording showed that E2 treatment decreased the peak IT and also reduced the low-threshold spikes (LTS) number, amplitude and width of LTS in the caudal LHb. Taken together, the results obtained in the present study suggest that E2 may inhibit T-channel activity by selectively down-regulating Cav3.3 calcium channel in the caudal LHb, leading to reduced the possibility of burst firing.

Spiking patterns of a neuron model to stimulus: Rich dynamics and oxygen's role.

Neuronal spiking patterns, which are of fundamental importance for the understanding of information processing in neural systems, can be generated in response to different stimuli. We here investigate in detail the stimulus-induced spiking patterns in a biologically plausible neuron model in which the oxygen concentration and the dynamical concentrations of potassium, sodium, and chloride are considered. Various types of spiking patterns can be induced by the different external potassium accumulations in response to the stimulus, including two different types of epileptic seizure (SZ) and spreading depression (SD) states, two different mixed states of SD and SZ, SZ state with multi-burst, and tonic firing behaviors. Interestingly, we show that these rich spiking patterns can also be induced by the current stimulus with a low oxygen concentration. Furthermore, we reveal that the stimulus can induce two different phase transitions from the SD state to the SZ state according to the phase transition theory, which results in the different electrical activities. All these findings may provide insight into information processing in neural systems.

Brainstem Steering of Locomotor Activity in the Newborn Rat.

Control of locomotion relies on motor loops conveying modulatory signals between brainstem and spinal motor circuits. We investigated the steering control of the brainstem reticular formation over the spinal locomotor networks using isolated brainstem-spinal cord preparations of male and female neonatal rats. First, we performed patch-clamp recordings of identified reticulospinal cells during episodes of fictive locomotion. This revealed that a spinal ascending phasic modulation of reticulospinal cell activity is already present at birth. Half of the cells exhibited tonic firing during locomotion, while the other half emitted phasic discharges of action potentials phase locked to ongoing activity. We next showed that mimicking the phasic activity of reticulospinal neurons by applying patterned electrical stimulation bilaterally at the ventral caudal medulla level triggered fictive locomotion efficiently. Moreover, the brainstem stimuli-induced locomotor rhythm was entrained in a one-to-one coupling over a range of cycle periods (2-6 s). Additionally, we induced turning like motor outputs by either increasing or decreasing the relative duration of the stimulation trains on one side of the brainstem compared to the other. The ability of the patterned descending command to control the locomotor output depended on the functional integrity of ventral reticulospinal pathways and the involvement of local spinal central pattern generator circuitry. Altogether, this study provides a mechanism by which brainstem reticulospinal neurons relay steering and speed commands to the spinal locomotor networks.SIGNIFICANCE STATEMENT Locomotor function allows the survival of most animal species while sustaining the expression of fundamental behaviors. Locomotor activities adapt from moment to moment to behavioral and environmental changes. We show that the brainstem can control the spinal locomotor network outputs through phasic descending commands that alternate bilaterally. Manipulating the periodicity and/or the relative durations of the left and right descending commands at the brainstem level is efficient to set the locomotor speed and sustain directional changes.

Low- and high-threshold primary afferent inputs to spinal lamina III antenna-type neurons.

The dorsal horn of the spinal cord (laminae I-VI) processes diverse modalities of nociceptive and nonnociceptive sensory information. Antenna-type neurons with cell bodies located in lamina III and large dendritic trees extending from the superficial lamina I to deep lamina IV are best shaped for the integration of a wide variety of inputs arising from primary afferent fibers and intrinsic spinal circuitries. Although the somatodendritic morphology, the hallmark of antenna neurons, has been well studied, little is still known about the axon structure and basic physiological properties of these cells. Here, we did whole-cell recordings in a rat (P9-P12) spinal cord preparation with attached dorsal roots to examine the axon course, intrinsic firing properties, and primary afferent inputs of antenna cells. Nine antenna cells were identified from a large sample of biocytin-filled lamina III neurons (n = 46). Axon of antenna cells showed intensive branching in laminae III-IV and, in half of the cases, issued dorsally directed collaterals reaching lamina I. Antenna cells exhibited tonic and rhythmic firing patterns; single spikes were followed by hyperpolarization or depolarization. The neurons received monosynaptic inputs from the low-threshold Aβ afferents, Aδ afferents, as well as from the high-threshold Aδ, and C afferents. When selectively activated, C-fiber-driven monosynaptic and polysynaptic excitatory postsynaptic potentials were sufficiently strong to evoke firing in the neurons. Thus, lamina III antenna neurons integrate low-threshold and nociceptive high-threshold primary afferent inputs and can function as wide dynamic range neurons able to directly connect deep dorsal horn with the major nociceptive projection area lamina I.

Thalamic dual control of sleep and wakefulness.

Slow-waves (0.5 - 4 Hz) predominate in the cortical electroencephalogram during non-rapid eye movement (NREM) sleep in mammals. They reflect the synchronization of large neuronal ensembles alternating between active (UP) and quiescent (Down) states and propagating along the neocortex. The thalamic contribution to cortical UP-states and sleep modulation remains unclear. Here we show that spontaneous firing of centromedial thalamus (CMT) neurons in mice is phase advanced to global cortical UP-states and NREM–wake transitions. Tonic optogenetic activation of CMT neurons induces NREM–wake transitions, whereas burst activation mimics UP-states in the cingulate cortex (CING) and enhances brain-wide synchrony of cortical slow-waves during sleep, through a relay in the antero-dorsal thalamus (AD). Finally, we demonstrate that CMT and AD relay neurons promote sleep recovery. These findings suggest that the firing pattern of CMT neurons can modulate brain-wide cortical activity during sleep and provides dual control of sleep-wake states.

Brain stimulation patterns emulating endogenous thalamocortical input to parvalbumin-expressing interneurons reduce nociception in mice

BackgroundThe bursting pattern of thalamocortical (TC) pathway dampens nociception. Whether brain stimulation mimicking endogenous patterns can engage similar sensory gating processes in the cortex and reduce nociceptive behaviors remains uninvestigated. ObjectiveWe investigated the role of cortical parvalbumin expressing (PV) interneurons within the TC circuit in gating nociception and their selective response to TC burst patterns. We then tested if transcranial magnetic stimulation (TMS) patterned on endogenous nociceptive TC bursting modulate nociceptive behaviors. MethodsThe switching of TC neurons between tonic (single spike) and burst (high frequency spikes) firing modes may be a critical component in modulating nociceptive signals. Deep brain electrical stimulation of TC neurons and immunohistochemistry were used to examine the differential influence of each firing mode on cortical PV interneuron activity. Optogenetic stimulation of cortical PV interneurons assessed a direct role in nociceptive modulation. A new TMS protocol mimicking thalamic burst firing patterns, contrasted with conventional continuous and intermittent theta burst protocols, tested if TMS patterned on endogenous TC activity reduces nociceptive behaviors in mice. ResultsImmunohistochemical evidence confirmed that burst, but not tonic, deep brain stimulation of TC neurons increased the activity of PV interneurons in the cortex. Both optogenetic activation of PV interneurons and TMS protocol mimicking thalamic burst reduced nociceptive behaviors. ConclusionsOur findings suggest that burst firing of TC neurons recruits PV interneurons in the cortex to reduce nociceptive behaviors and that neuromodulation mimicking thalamic burst firing may be useful for modulating nociception.

All-Optical Electrophysiology for High-Throughput Functional Characterization of a Human iPSC-Derived Motor Neuron Model of ALS.

SummaryHuman induced pluripotent stem cell (iPSC)-derived neurons are an attractive substrate for modeling disease, yet the heterogeneity of these cultures presents a challenge for functional characterization by manual patch-clamp electrophysiology. Here, we describe an optimized all-optical electrophysiology, “Optopatch,” pipeline for high-throughput functional characterization of human iPSC-derived neuronal cultures. We demonstrate the method in a human iPSC-derived motor neuron (iPSC-MN) model of amyotrophic lateral sclerosis (ALS). In a comparison of iPSC-MNs with an ALS-causing mutation (SOD1 A4V) with their genome-corrected controls, the mutants showed elevated spike rates under weak or no stimulus and greater likelihood of entering depolarization block under strong optogenetic stimulus. We compared these results with numerical simulations of simple conductance-based neuronal models and with literature results in this and other iPSC-based models of ALS. Our data and simulations suggest that deficits in slowly activating potassium channels may underlie the changes in electrophysiology in the SOD1 A4V mutation.

Inhibition of striatal cholinergic interneuron activity by the Kv7 opener retigabine and the nonsteroidal anti-inflammatory drug diclofenac

Striatal cholinergic interneurons provide modulation to striatal circuits involved in voluntary motor control and goal-directed behaviors through their autonomous tonic discharge and their firing "pause" responses to novel and rewarding environmental events. Striatal cholinergic interneuron hyperactivity was linked to the motor deficits associated with Parkinson's disease and the adverse effects of chronic antiparkinsonian therapy like l-DOPA-induced dyskinesia. Here we addressed whether Kv7 channels, which provide negative feedback to excitation in other neuron types, are involved in the control of striatal cholinergic interneuron tonic activity and response to excitatory inputs. We found that autonomous firing of striatal cholinergic interneurons is not regulated by Kv7 channels. In contrast, Kv7 channels limit the summation of excitatory postsynaptic potentials in cholinergic interneurons through a postsynaptic mechanism. Striatal cholinergic interneurons have a high reserve of Kv7 channels, as their opening using pharmacological tools completely silenced the tonic firing and markedly reduced their intrinsic excitability. A strong inhibition of striatal cholinergic interneurons was also observed in response to the anti-inflammatory drugs diclofenac and meclofenamic acid, however, this effect was independent of Kv7 channels. These data bring attention to new potential molecular targets and pharmacological tools to control striatal cholinergic interneuron activity in pathological conditions where they are believed to be hyperactive, including Parkinson's disease.

Differential inputs to striatal cholinergic and parvalbumin interneurons imply functional distinctions.

Striatal cholinergic (ChAT) and parvalbumin (PV) interneurons exert powerful influences on striatal function in health and disease, yet little is known about the organization of their inputs. Here using rabies tracing, electrophysiology and genetic tools, we compare the whole-brain inputs to these two types of striatal interneurons and dissect their functional connectivity in mice. ChAT interneurons receive a substantial cortical input from associative regions of cortex, such as the orbitofrontal cortex. Amongst subcortical inputs, a previously unknown inhibitory thalamic reticular nucleus input to striatal PV interneurons is identified. Additionally, the external segment of the globus pallidus targets striatal ChAT interneurons, which is sufficient to inhibit tonic ChAT interneuron firing. Finally, we describe a novel excitatory pathway from the pedunculopontine nucleus that innervates ChAT interneurons. These results establish the brain-wide direct inputs of two major types of striatal interneurons and allude to distinct roles in regulating striatal activity and controlling behavior.

(Invited) Imaging Dopamine Neuromodulation with Single Wall Carbon Nanotube Sensors

Neurons communicate through chemical neurotransmitter signals that either terminate at the postsynaptic process (“wired transmission”) or diffuse beyond the synaptic cleft to modulate the activity of larger neuronal networks (“volume transmission”). Molecules such as dopamine belong to the latter class of neurotransmitters, for which real-time imaging of the signal’s spatial propagation would constitute a major advance in neurochemical imaging. To this end, we present a nanoscale near-infrared fluorescent sensor for dopamine and demonstrate its efficacy for imaging dopamine volume transmission in the extracellular space of the brain striatum and prefrontal cortex. The sensor is developed from polymers pinned to the surface of single wall carbon nanotubes (SWNT) in which the surface-adsorbed polymer is the recognition moiety and the carbon nanotube the fluorescence transduction element. Using this paradigm, we describe the design, characterization, and implementation of polymer-functionalized SWNT nanosensors and present recent progress in imaging the dynamics of evoked and spontaneous dopamine release in mouse acute brain slices. Fluorescence imaging of dopamine volume transmission with our probes provides much needed spatial information and high resolution of dopamine dynamics in contrast with traditional electrochemical dopamine measurements obtained with fast scan cyclic voltammetry (FSCV). Whereas FSCV measures dopamine concentration at a single point arising from the activity of a large number of dopamine terminals, our probes capture local release arising from single and multiple terminals within the imaging field of view. From imaging data, we can describe localized release and clearance kinetics of dopamine neuromodulation. In the brain striatum, the nanosensors record spatially and temporally resolved dynamic behavior of dopamine on length scales that encompass single terminals and inter-terminal distances, and enable investigation of the effect of antidepressant drugs such as Nomifensine on dopamine dynamics. In the prefrontal cortex (PFC), an area of sparse dopaminergic innervation, dopamine nanosensors capture dynamics elicited by single terminals. In both the striatum and PFC, optogenetics enables selective stimulation of dopaminergic terminals to evoke dopamine release. We further describe how nanoparticle exciton engineering can be used to tune sensor performance and design sensors best suited for in vivo experimentation. Molecular dynamics modeling elucidates the physicochemical phenomena underlying the dopamine sensing process and stochastic simulations reveal that optical nanosensors can capture millisecond-scale changes during phasic and tonic firing of dopaminergic neurons. Combining our experimental and computational work, we outline the functional range of this technology for imaging dopamine neuromodulation in biologically complex and optically dense live brain. Our experimental and theoretical results show that near-infrared (nIR) neurotransmitter nanosensor constructs can relay information about neuronal signaling in the tissue-compatible nIR optical window, with spatiotemporal scales that capture both single synaptic release events and ensemble terminal behavior suitable for in vivo behavioral experimentation.

Neural representations of time-linked memory.

Many cognitive processes, such as episodic memory and decision making, rely on the ability to form associations between two events that occur separately in time. The formation of such temporal associations depends on neural representations of three types of information: what has been presented (trace holding), what will follow (temporal expectation), and when the following event will occur (explicit timing). The present review seeks to link these representations with firing patterns of single neurons recorded while rodents and non-human primates associate stimuli, outcomes, and motor responses over time intervals. Across these studies, two distinct firing patterns were observed in the hippocampus, neocortex, and striatum: some neurons change firing rates during or shortly after the stimulus presentation and sustain the firing rate stably or sidlingly during the subsequent intervals (tonic firings). Other neurons transiently change firing rates during a specific moment within the time intervals (phasic firings), and as a group, they form a sequential firing pattern that covers the entire interval. Clever task designs used in some of these studies collectively provide evidence that both tonic and phasic firing responses represent trace holding, temporal expectation, and explicit timing. Subsequently, we applied machine-learning based classification approaches to the two firing patterns within the same dataset collected from rat medial prefrontal cortex during trace eyeblink conditioning. This quantitative analysis revealed that phasic-firing patterns showed greater selectivity for stimulus identity and temporal position than tonic-firing patterns. Our summary illuminates distributed neural representations of temporal association in the forebrain and generates several ideas for future investigations.

Pauses in cholinergic interneuron firing exert an inhibitory control on striatal output in vivo.

The cholinergic interneurons (CINs) of the striatum are crucial for normal motor and behavioral functions of the basal ganglia. Striatal CINs exhibit tonic firing punctuated by distinct pauses. Pauses occur in response to motivationally significant events, but their function is unknown. Here we investigated the effects of pauses in CIN firing on spiny projection neurons (SPNs) - the output neurons of the striatum - using in vivo whole cell and juxtacellular recordings in mice. We found that optogenetically-induced pauses in CIN firing inhibited subthreshold membrane potential activity and decreased firing of SPNs. During pauses, SPN membrane potential fluctuations became more hyperpolarized and UP state durations became shorter. In addition, short-term plasticity of corticostriatal inputs was decreased during pauses. Our results indicate that, in vivo, the net effect of the pause in CIN firing on SPNs activity is inhibition and provide a novel mechanism for cholinergic control of striatal output.

Characterization of Superficial Dorsal Horn Neurons from “Tamamaki” Mice and Stability of their GAD67-EGFP Phenotype in Defined-Medium Organotypic Culture

Defined medium organotypic cultures (DMOTC) containing spinal dorsal horn neurons are especially useful in studying the etiology and pharmacology of chronic pain. We made whole-cell recordings from neurons in acutely isolated mouse spinal cord slices or from those maintained in DMOTC for up to 6 weeks. In acute slices, neurons in the substantia gelatinosa exhibited 7 different firing patterns in response to 800-ms depolarizing current commands; delay (irregular), delay (tonic), tonic, regular firing, phasic, initial bursting and single spiking. Initial bursting and regular firing neurons are not found in rat substantia gelatinosa. In acute slices from “Tamamaki” mice that express enhanced green fluorescent protein (EGFP) under the control of the glutamic acid decarboxylase 67 (GAD67) promotor, tonic, phasic and regular firing neurons exhibited the strongest GABAergic (GAD67-EGFP+) phenotype. Delay (tonic) and delay (irregular) neurons almost never expressed GAD67 (GAD67-EGFP−) and are likely glutamatergic. All seven phenotypes were preserved in mouse spinal cord neurons in DMOTC prepared from e12 embryos and the GAD67-EGFP+ phenotype continued to associate with phasic and regular firing neurons. Only 3 out of 51 GAD67-EGFP+ neurons exhibited a delay (tonic) firing pattern. Modifications to the mouse genome thus continue to be expressed when embryonic neurons develop in vitro in DMOTC. However, analysis of the amplitude and interevent interval of spontaneous EPSCs (sEPSCs) indicated substantial re-arrangement of synaptic connections within the cultures. Despite this, the characteristics and age-dependence of asynchronous oscillatory activity, as monitored by multiphoton Ca2+ imaging, were similar in acute slices and in DMOTC.

Role of the Axon Initial Segment in the Control of Spontaneous Frequency of Nigral Dopaminergic Neurons In Vivo.

The spontaneous tonic discharge activity of nigral dopamine neurons plays a fundamental role in dopaminergic signaling. To investigate the role of neuronal morphology and architecture with respect to spontaneous activity in this population, we visualized the 3D structure of the axon initial segment (AIS) along with the entire somatodendritic domain of adult male mouse dopaminergic neurons, previously recorded in vivo We observed a positive correlation of the firing rate with both proximity and size of the AIS. Computational modeling showed that the size of the AIS, but not its position within the somatodendritic domain, is the major causal determinant of the tonic firing rate in the intact model, by virtue of the higher intrinsic frequency of the isolated AIS. Further mechanistic analysis of the relationship between neuronal morphology and firing rate showed that dopaminergic neurons function as a coupled oscillator whose frequency of discharge results from a compromise between AIS and somatodendritic oscillators. Thus, morphology plays a critical role in setting the basal tonic firing rate, which in turn could control striatal dopaminergic signaling that mediates motivation and movement.SIGNIFICANCE STATEMENT The frequency at which nigral dopamine neurons discharge action potentials sets baseline dopamine levels in the brain, which enables activity in motor, cognitive, and motivational systems. Here, we demonstrate that the size of the axon initial segment, a subcellular compartment responsible for initiating action potentials, is a key determinant of the firing rate in these neurons. The axon initial segment and all the molecular components that underlie its critical function may provide a novel target for the regulation of dopamine levels in the brain.