D2 dopamine receptors and the striatopallidal pathway modulate L-DOPA-induced dyskinesia in the mouse.

L-DOPA-induced dyskinesia (LID) remains a major complication of Parkinson's disease management for which better therapies are necessary. The contribution of the striatonigral direct pathway to LID is widely acknowledged but whether the striatopallidal pathway is involved remains debated. Selective optogenetic stimulation of striatonigral axon terminals induces dyskinesia in mice rendered hemiparkinsonian with the toxin 6-hydroxydopamine (6-OHDA). Here we show that optogenetically-induced dyskinesia is increased by the D2-type dopamine receptor agonist quinpirole. Although the quinpirole effect may be mediated by D2 receptor stimulation in striatopallidal neurons, alternative mechanisms may be responsible as well. To selectively modulate the striatopallidal pathway, we selectively expressed channelrhodopsin-2 (ChR2) in D2 receptor expressing neurons by crossing D2-Cre and ChR2-flox mice. The animals were rendered hemiparkinsonian and implanted with an optic fiber at the ipsilateral external globus pallidus (GPe). Stimulation of ChR2 at striatopallidal axon terminals reduced LID and also general motility during the off L-DOPA state, without modifying the pro-motor effect of low doses of L-DOPA producing mild or no dyskinesia. Overall, the present study shows that D2-type dopamine receptors and the striatopallidal pathway modulate dyskinesia and suggest that targeting striatopallidal axon terminals at the GPe may have therapeutic potential in the management of LID.

Modulation of cortical slow oscillatory rhythm by GABAB receptors: an in vitro experimental and computational study.

KEY POINTS: We confirm that GABAB receptors (GABAB -Rs) are involved in the termination of Up-states; their blockade consistently elongates Up-states. GABAB -Rs also modulate Down-states and the oscillatory cycle, thus having an impact on slow oscillation rhythm and its regularity. The most frequent effect of GABAB -R blockade is elongation of Down-states and subsequent decrease of oscillatory frequency, with an increased regularity. In a quarter of cases, GABAB -R blockade shortened Down-states and increased oscillatory frequency, changes that are independent of firing rates in Up-states. Our computer model provides mechanisms for the experimentally observed dynamics following blockade of GABAB -Rs, for Up/Down durations, oscillatory frequency and regularity. The time course of excitation, inhibition and adaptation can explain the observed dynamics of the network. This study brings novel insights into the role of GABAB -R-mediated slow inhibition on the slow oscillatory activity, which is considered the default activity pattern of the cortical network. ABSTRACT: Slow wave oscillations (SWOs) dominate cortical activity during deep sleep, anaesthesia and in some brain lesions. SWOs are composed of periods of activity (Up states) interspersed with periods of silence (Down states). The rhythmicity expressed during SWOs integrates neuronal and connectivity properties of the network and is often altered under pathological conditions. Adaptation mechanisms as well as synaptic inhibition mediated by GABAB receptors (GABAB -Rs) have been proposed as mechanisms governing the termination of Up states. The interplay between these two mechanisms is not well understood, and the role of GABAB -Rs controlling the whole cycle of the SWO has not been described. Here we contribute to its understanding by combining in vitro experiments on spontaneously active cortical slices and computational techniques. GABAB -R blockade modified the whole SWO cycle, not only elongating Up states, but also affecting the subsequent Down state duration. Furthermore, while adaptation tends to yield a rather regular behaviour, we demonstrate that GABAB -R activation desynchronizes the SWOs. Interestingly, variability changes could be accomplished in two different ways: by either shortening or lengthening the duration of Down states. Even when the most common observation following GABAB -Rs blocking is the lengthening of Down states, both changes are expressed experimentally and also in numerical simulations. Our simulations suggest that the sluggishness of GABAB -Rs to follow the excitatory fluctuations of the cortical network can explain these different network dynamics modulated by GABAB -Rs.

Direct pathway neurons in mouse dorsolateral striatum in vivo receive stronger synaptic input than indirect pathway neurons.

Striatal projection neurons, the medium spiny neurons (MSNs), play a crucial role in various motor and cognitive functions. MSNs express either D1- or D2-type dopamine receptors and initiate the direct-pathway (dMSNs) or indirect pathways (iMSNs) of the basal ganglia, respectively. dMSNs have been shown to receive more inhibition than iMSNs from intrastriatal sources. Based on these findings, computational modeling of the striatal network has predicted that under healthy conditions dMSNs should receive more total input than iMSNs. To test this prediction, we analyzed in vivo whole cell recordings from dMSNs and iMSNs in healthy and dopamine-depleted (6OHDA) anaesthetized mice. By comparing their membrane potential fluctuations, we found that dMSNs exhibited considerably larger membrane potential fluctuations over a wide frequency range. Furthermore, by comparing the spike-triggered average membrane potentials, we found that dMSNs depolarized toward the spike threshold significantly faster than iMSNs did. Together, these findings (in particular the STA analysis) corroborate the theoretical prediction that direct-pathway MSNs receive stronger total input than indirect-pathway neurons. Finally, we found that dopamine-depleted mice exhibited no difference between the membrane potential fluctuations of dMSNs and iMSNs. These data provide new insights into the question of how the lack of dopamine may lead to behavioral deficits associated with Parkinson's disease.NEW & NOTEWORTHY The direct and indirect pathways of the basal ganglia originate from the D1- and D2-type dopamine receptor expressing medium spiny neurons (dMSNs and iMSNs). Theoretical results have predicted that dMSNs should receive stronger synaptic input than iMSNs. Using in vivo intracellular membrane potential data, we provide evidence that dMSNs indeed receive stronger input than iMSNs, as has been predicted by the computational model.

Distinct Corticostriatal and Intracortical Pathways Mediate Bilateral Sensory Responses in the Striatum.

Individual striatal neurons integrate somatosensory information from both sides of the body, however, the afferent pathways mediating these bilateral responses are unclear. Whereas ipsilateral corticostriatal projections are prevalent throughout the neocortex, contralateral projections provide sparse input from primary sensory cortices, in contrast to the dense innervation from motor and frontal regions. There is, therefore, an apparent discrepancy between the observed anatomical pathways and the recorded striatal responses. We used simultaneous in vivo whole-cell and extracellular recordings combined with focal cortical silencing, to dissect the afferent pathways underlying bilateral sensory integration in the mouse striatum. We show that unlike direct corticostriatal projections mediating responses to contralateral whisker deflection, responses to ipsilateral stimuli are mediated mainly by intracortical projections from the contralateral somatosensory cortex (S1). The dominant pathway is the callosal projection from contralateral to ipsilateral S1. Our results suggest a functional difference between the cortico-basal ganglia pathways underlying bilateral sensory and motor processes.

Gain modulation of synaptic inputs by network state in auditory cortex in vivo.

The cortical network recurrent circuitry generates spontaneous activity organized into Up (active) and Down (quiescent) states during slow-wave sleep or anesthesia. These different states of cortical activation gain modulate synaptic transmission. However, the reported modulation that Up states impose on synaptic inputs is disparate in the literature, including both increases and decreases of responsiveness. Here, we tested the hypothesis that such disparate observations may depend on the intensity of the stimulation. By means of intracellular recordings, we studied synaptic transmission during Up and Down states in rat auditory cortex in vivo. Synaptic potentials were evoked either by auditory or electrical (thalamocortical, intracortical) stimulation while randomly varying the intensity of the stimulus. Synaptic potentials evoked by the same stimulus intensity were compared in Up/Down states. Up states had a scaling effect on the stimulus-evoked synaptic responses: the amplitude of weaker responses was potentiated whereas that of larger responses was maintained or decreased with respect to the amplitude during Down states. We used a computational model to explore the potential mechanisms explaining this nontrivial stimulus-response relationship. During Up/Down states, there is different excitability in the network and the neuronal conductance varies. We demonstrate that the competition between presynaptic recruitment and the changing conductance might be the central mechanism explaining the experimentally observed stimulus-response relationships. We conclude that the effect that cortical network activation has on synaptic transmission is not constant but contingent on the strength of the stimulation, with a larger modulation for stimuli involving both thalamic and cortical networks.

Spinal glutamatergic neurons defined by EphA4 signaling are essential components of normal locomotor circuits.

EphA4 signaling is essential for the spatiotemporal organization of neuronal circuit formation. In mice, deletion of this signaling pathway causes aberrant midline crossing of axons from both brain and spinal neurons and the complete knock-outs (KOs) exhibit a pronounced change in motor behavior, where alternating gaits are replaced by a rabbit-like hopping gait. The neuronal mechanism that is responsible for the gait switch in these KO mice is not known. Here, using intersectional genetics, we demonstrate that a spinal cord-specific deletion of EphA4 signaling is sufficient to generate the overground hopping gait. In contrast, selective deletion of EphA4 signaling in forebrain neurons, including the corticospinal tract neurons, did not result in a change in locomotor pattern. The gait switch was attributed to the loss of EphA4 signaling in excitatory Vglut2+ neurons, which is accompanied by an increased midline crossing of Vglut2+ neurons in the ventral spinal cord. Our findings functionally define spinal EphA4 signaling in excitatory Vglut2+ neurons as required for proper organization of the spinal locomotor circuitry, and place these cells as essential components of the mammalian locomotor network.

Callosal inputs generate side-invariant receptive fields in the barrel cortex.

Barrel cortex integrates contra- and ipsilateral whiskers' inputs. While contralateral inputs depend on the thalamocortical innervation, ipsilateral ones are thought to rely on callosal axons. These are more abundant in the barrel cortex region bordering with S2 and containing the row A-whiskers representation, the row lying nearest to the facial midline. Here, we ask what role this callosal axonal arrangement plays in ipsilateral tactile signaling. We found that novel object exploration with ipsilateral whiskers confines c-Fos expression within the highly callosal subregion. Targeting this area with in vivo patch-clamp recordings revealed neurons with uniquely strong ipsilateral responses dependent on the corpus callosum, as assessed by tetrodotoxin silencing and by optogenetic activation of the contralateral hemisphere. Still, in this area, stimulation of contra- or ipsilateral row A-whiskers evoked an indistinguishable response in some neurons, mostly located in layers 5/6, indicating their involvement in the midline representation of the whiskers' sensory space.

Medium spiny neurons activity reveals the discrete segregation of mouse dorsal striatum.

Behavioral studies differentiate the rodent dorsal striatum (DS) into lateral and medial regions; however, anatomical evidence suggests that it is a unified structure. To understand striatal dynamics and basal ganglia functions, it is essential to clarify the circuitry that supports this behavioral-based segregation. Here, we show that the mouse DS is made of two non-overlapping functional circuits divided by a boundary. Combining in vivo optopatch-clamp and extracellular recordings of spontaneous and evoked sensory activity, we demonstrate different coupling of lateral and medial striatum to the cortex together with an independent integration of the spontaneous activity, due to particular corticostriatal connectivity and local attributes of each region. Additionally, we show differences in slow and fast oscillations and in the electrophysiological properties between striatonigral and striatopallidal neurons. In summary, these results demonstrate that the rodent DS is segregated in two neuronal circuits, in homology with the caudate and putamen nuclei of primates.

GABAB receptors: modulation of thalamocortical dynamics and synaptic plasticity.

GABAB-receptors (GABAB-Rs) are metabotropic, G protein-coupled receptors for the neurotransmitter GABA. Their activation induces slow inhibitory control of the neuronal excitability mediated by pre- and postsynaptic inhibition. Presynaptically GABAB-Rs reduce GABA and glutamate release inhibiting presynaptic Ca2+ channels in both inhibitory and excitatory synapses while postsynaptic GABAB-Rs induce robust slow hyperpolarization by the activation of K+ channels. GABAB-Rs are activated by non-synaptic or volume transmission, which requires high levels of GABA release, either by the simultaneous discharge of GABAergic interneurons or very intense discharges in the thalamus or by means of the activation of a neurogliaform interneurons in the cortex. The main receptor subunits GABAB1a, GABAB1b and GABAB2 are strongly expressed in neurons and glial cells throughout the central nervous system and GABAB-R activation is related to many neuronal processes such as the modulation of rhythmic activity in several brain regions. In the thalamus, GABAB-Rs modulate the generation of the main thalamic rhythm, spindle waves. In the cerebral cortex, GABAB-Rs also modulate the most prominent emergent oscillatory activity-slow oscillations-as well as faster oscillations like gamma frequency. Further, recent studies evaluating the complexity expressed by the cortical network, a parameter associated with consciousness levels, have found that GABAB-Rs enhance this complexity, while their blockade decreases it. This review summarizes the current results on how the activation of GABAB-Rs affects the interchange of information between brain areas by controlling rhythmicity as well as synaptic plasticity.

Inhibitory modulation of cortical up states.

The balance between excitation and inhibition is critical in the physiology of the cerebral cortex. To understand the influence of inhibitory control on the emergent activity of the cortical network, inhibition was progressively blocked in a slice preparation that generates spontaneous rhythmic up states at a similar frequency to those occurring in vivo during slow-wave sleep or anesthesia. Progressive removal of inhibition induced a parametric shortening of up state duration and elongation of the down states, the frequency of oscillations decaying. Concurrently, a gradual increase in the network firing rate during up states occurred. The slope of transitions between up and down states was quantified for different levels of inhibition. The slope of upward transitions reflects the recruitment of the local network and was progressively increased when inhibition was decreased, whereas the speed of activity propagation became faster. Removal of inhibition eventually resulted in epileptiform activity. Whereas gradual reduction of inhibition induced linear changes in up/down states and their propagation, epileptiform activity was the result of a nonlinear transformation. A computational network model showed that strong recurrence plus activity-dependent hyperpolarizing currents were sufficient to account for the observed up state modulations and predicted an increase in activity-dependent hyperpolarization following up states when inhibition was decreased, which was confirmed experimentally.

Effective reduced diffusion-models: a data driven approach to the analysis of neuronal dynamics.

We introduce in this paper a new method for reducing neurodynamical data to an effective diffusion equation, either experimentally or using simulations of biophysically detailed models. The dimensionality of the data is first reduced to the first principal component, and then fitted by the stationary solution of a mean-field-like one-dimensional Langevin equation, which describes the motion of a Brownian particle in a potential. The advantage of such description is that the stationary probability density of the dynamical variable can be easily derived. We applied this method to the analysis of cortical network dynamics during up and down states in an anesthetized animal. During deep anesthesia, intracellularly recorded up and down states transitions occurred with high regularity and could not be adequately described by a one-dimensional diffusion equation. Under lighter anesthesia, however, the distributions of the times spent in the up and down states were better fitted by such a model, suggesting a role for noise in determining the time spent in a particular state.

Spontaneous high-frequency (10-80 Hz) oscillations during up states in the cerebral cortex in vitro.

High-frequency oscillations in cortical networks have been linked to a variety of cognitive and perceptual processes. They have also been recorded in small cortical slices in vitro, indicating that neuronal synchronization at these frequencies is generated in the local cortical circuit. However, in vitro experiments have hitherto necessitated exogenous pharmacological or electrical stimulation to generate robust synchronized activity in the beta/gamma range. Here, we demonstrate that the isolated cortical microcircuitry generates beta and gamma oscillations spontaneously in the absence of externally applied neuromodulators or synaptic agonists. We show this in a spontaneously active slice preparation that engages in slow oscillatory activity similar to activity during slow-wave sleep. beta and gamma synchronization appeared during the up states of the slow oscillation. Simultaneous intracellular and extracellular recordings revealed synchronization between the timing of incoming synaptic events and population activity. This rhythm was mechanistically similar to pharmacologically induced gamma rhythms, as it also included sparse, irregular firing of neurons within the population oscillation, predominant involvement of inhibitory neurons, and a decrease of oscillation frequency after barbiturate application. Finally, we show in a computer model how a synaptic loop between excitatory and inhibitory neurons can explain the emergence of both the slow (<1 Hz) and the beta-range oscillations in the neocortical network. We therefore conclude that oscillations in the beta/gamma range that share mechanisms with activity reported in vivo or in pharmacologically activated in vitro preparations can be generated during slow oscillatory activity in the local cortical circuit, even without exogenous pharmacological or electrical stimulation.

A new paradigm for the reversible blockage of whisker sensory transmission.

The objective of this study was to explore a paradigm that would allow a temporary deprivation of whisker information lasting for a few hours. An additional requirement was to be non-invasive in order to be usable in awake chronically implanted rats without inducing stress. With that aim, electrophysiological recordings from the barrel cortex of anesthetized rats were obtained. The pressure of an air-puff (5-10 ms) delivered to the whiskers was adjusted to evoke a consistent response of around 100 microV (extracellular) or approximately 5 mV (intracellular) in the contralateral cortex. Lidocaine was then locally applied in different forms (cream, local injection, aerosol, drops) and concentrations (2-10%) to the base of the whiskers. The stimulus-induced response was monitored once every 5s for several hours (3-6h) in order to characterize its course of action. Local injection of lidocaine induced the fastest and most complete blockage, but was ruled out for being invasive. Out of the remaining forms of application, a lidocaine drop (0.4 ml, 10%) to the base of the whiskers was found to induce a reliable blockage (to an average 9% the original response). The maximum effect was reached after 150-200 min, and the response was totally recovered approximately 300 min after lidocaine application. This characterization should be useful to induce an efficient, short term and reversible blockage of whisker sensory transmission in both anesthetized and awake preparations, while not causing stress in an awake animal.

A New Micro-holder Device for Local Drug Delivery during In Vivo Whole-cell Recordings.

Focal administration of pharmacological agents during in vivo recordings is a useful technique to study the functional properties of neural microcircuits. However, the lack of visual control makes this task difficult and inaccurate, especially when targeting small and deep regions where spillover to neighboring regions is likely to occur. An additional problem with recording stability arises when combining focal drug administration with in vivo intracellular recordings, which are highly sensitive to mechanical vibrations. To address these technical issues, we designed a micro-holder that enables accurate local application of pharmacological agents during in vivo whole-cell recordings. The holder couples the recording and drug delivery pipettes with adjustable distance between the respective tips adapted to the experimental needs. To test the efficacy of the micro-holder we first performed whole-cell recordings in mouse primary somatosensory cortex (S1) with simultaneous extracellular recordings in S1 and motor cortex (M1), before and after local application of bicuculline methiodide (BMI 200 µM). The blockade of synaptic inhibition resulted in increased amplitudes and rising slopes of "Up states", and shortening of their duration. We then checked the usability of the micro-holder in a deeper brain structure, the striatum. We applied tetrodotoxin (TTX 10 µM) during whole-cell recordings in the striatum, while simultaneously obtaining extracellular recordings in S1 and M1. The focal application of TTX in the striatum blocked Up states in the recorded striatal neurons, without affecting the cortical activity. We also describe two different approaches for precisely releasing the drugs without unwanted leakage along the pipette approach trajectory.

Multisensory integration in the mouse striatum.

The basal ganglia are involved in sensorimotor functions and action selection, both of which require the integration of sensory information. In order to determine how such sensory inputs are integrated, we obtained whole-cell recordings in mouse dorsal striatum during presentation of tactile and visual stimuli. All recorded neurons responded to bilateral whisker stimulation, and a subpopulation also responded to visual stimulation. Neurons responding to both visual and tactile stimuli were located in dorsomedial striatum, whereas those responding only to whisker deflections were located dorsolaterally. Responses were mediated by overlapping excitation and inhibition, with excitation onset preceding that of inhibition by several milliseconds. Responses differed according to the type of neuron, with direct pathway MSNs having larger responses and longer latencies between ipsilateral and contralateral responses than indirect pathway MSNs. Our results suggest that striatum acts as a sensory "hub" with specialized functional roles for the different neuron types.

Synaptic transmission and plasticity in an active cortical network.

BACKGROUND: The cerebral cortex is permanently active during both awake and sleep states. This ongoing cortical activity has an impact on synaptic transmission and short-term plasticity. An activity pattern generated by the cortical network is a slow rhythmic activity that alternates up (active) and down (silent) states, a pattern occurring during slow wave sleep, anesthesia and even in vitro. Here we have studied 1) how network activity affects short term synaptic plasticity and, 2) how synaptic transmission varies in up versus down states. METHODOLOGY/PRINCIPAL FINDINGS: Intracellular recordings obtained from cortex in vitro and in vivo were used to record synaptic potentials, while presynaptic activation was achieved either with electrical or natural stimulation. Repetitive activation of layer 4 to layer 2/3 synaptic connections from ferret visual cortex slices displayed synaptic augmentation that was larger and longer lasting in active than in silent slices. Paired-pulse facilitation was also significantly larger in an active network and it persisted for longer intervals (up to 200 ms) than in silent slices. Intracortical synaptic potentials occurring during up states in vitro increased their amplitude while paired-pulse facilitation disappeared. Both intracortical and thalamocortical synaptic potentials were also significantly larger in up than in down states in the cat visual cortex in vivo. These enhanced synaptic potentials did not further facilitate when pairs of stimuli were given, thus paired-pulse facilitation during up states in vivo was virtually absent. Visually induced synaptic responses displayed larger amplitudes when occurring during up versus down states. This was further tested in rat barrel cortex, where a sensory activated synaptic potential was also larger in up states. CONCLUSIONS/SIGNIFICANCE: These results imply that synaptic transmission in an active cortical network is more secure and efficient due to larger amplitude of synaptic potentials and lesser short term plasticity.

Impact of cortical network activity on short-term synaptic depression.

Repetitive stimulation of synaptic connections in the cerebral cortex often induces short-term synaptic depression (STD), a property directly related to the probability of transmitter release and critical for the computational properties of the network. In order to explore how spontaneous activity in the network affects this property, we first studied STD in cortical slices that were either silent or that displayed spontaneous rhythmic slow oscillations resembling those recorded during slow wave sleep in vivo. STD was considerably reduced by the occurrence of spontaneous rhythmic activity in the cortical network. Once the rhythmic activity started, depression decreased over time in parallel with the duration and intensity of the ongoing activity until a plateau was reached. Thalamocortical and intracortical synaptic potentials studied in vivo also showed stronger depression in a silent than in an active cortical network, and the depression values in the active cortical network in vivo were indistinguishable from those found in active slices in vitro. We suggest that this phenomenon is due to the different steady states of the synapses in active and in silent networks.