The mammalian motor system is organized around distinct sub-cortical subsystems, suggesting that intracortical circuits immediately upstream of spinal cord and basal ganglia might be functionally differentiated, too. Here, we show that the main excitatory pathway within mouse motor cortex, layer 2/3→5, is fractionated into distinct pathways targeting corticospinal and corticostriatal neurons, key cell classes involved in motor control. However, connections were selective for neurons in certain sub-layers: corticospinal neurons in upper layer 5B, and corticostriatal neurons in lower 5A. A simple structural combinatorial principle accounts for this highly specific functional circuit architecture: potential connectivity is established by neuronal sub-layer positioning, and actual connectivity within this framework is determined by long-range axonal projection targets. Thus, intracortical circuits of these pyramidal neurons are specified not only by their long-range axonal targets, or their layer or sub-layer positions, but by both, in specific combinations.

The generation of purposive movement by mammals involves coordinated activity in the corticospinal and corticostriatal systems, which are involved in different aspects of motor control. In the motor cortex, corticospinal and corticostriatal neurons are closely intermingled, raising the question of whether and how information flows intracortically within and across these two channels. To explore this, we developed an optogenetic technique based on retrograde transfection of neurons with deletion-mutant rabies virus encoding channelrhodopsin-2 (RV-ChR2), and used this in conjunction with retrograde anatomical labeling to stimulate and record from identified projection neurons in mouse motor cortex. We also used paired recordings to measure unitary connections. Both corticospinal and callosally projecting corticostriatal neurons in layer 5B formed within-class (recurrent) connections, with higher connection probability among corticostriatal than among corticospinal neurons. In contrast, across-class connectivity was extraordinarily asymmetric, essentially unidirectional from corticostriatal to corticospinal. Corticostriatal neurons in layer 5A and corticocortical neurons (callosal projection neurons similar to corticostriatal neurons) similarly received a paucity of corticospinal input. Connections involving presynaptic corticostriatal neurons had greater synaptic depression, and those involving postsynaptic corticospinal neurons had faster decaying EPSPs. Consequently, the three connections displayed a diversity of dynamic properties reflecting the different combinations of pre- and postsynaptic projection neurons. Collectively, these findings delineate a four-way specialized excitatory microcircuit formed by corticospinal and corticostriatal neurons. The “rectifying” corticostriatal-to-corticospinal connectivity implies a hierarchical organization and functional compartmentalization of corticospinal activity via unidirectional signaling from higher-order (corticostriatal) to lower order (corticospinal) output neurons.

Tinnitus has been associated with increased spontaneous and evoked activity, increased neural synchrony, and reorganization of tonotopic maps of auditory nuclei. However, the neurotransmitter systems mediating these changes are poorly understood. Here, we developed an in vitro assay that allows us to evaluate the roles of excitation and inhibition in determining the neural correlates of tinnitus. To measure the magnitude and spatial spread of evoked circuit activity, we used flavoprotein autofluorescence (FA) imaging, a metabolic indicator of neuronal activity. We measured FA responses after electrical stimulation of glutamatergic axons in slices containing the dorsal cochlear nucleus, an auditory brainstem nucleus hypothesized to be crucial in the triggering and modulation of tinnitus. FA imaging in dorsal cochlear nucleus brain slices from mice with behavioral evidence of tinnitus (tinnitus mice) revealed enhanced evoked FA response at the site of stimulation and enhanced spatial propagation of FA response to surrounding sites. Blockers of GABAergic inhibition enhanced FA response to a greater extent in control mice than in tinnitus mice. Blockers of excitation decreased FA response to a similar extent in tinnitus and control mice. These findings indicate that auditory circuits in mice with behavioral evidence of tinnitus respond to stimuli in a more robust and spatially distributed manner because of a decrease in GABAergic inhibition.T innitus, the persistent perception of a subjective sound in the absence of an acoustic stimulus (ringing of the ears), is often a debilitating condition that reduces the quality of life for many of those chronically affected. Estimates of the number of people experiencing tinnitus range from 8% to 20% of the general population (1). Despite the prevalence and growing incidence of tinnitus, the mechanisms underlying the induction and maintenance of tinnitus remain poorly understood.Animal models of tinnitus have contributed significantly to the understanding of the pathophysiology of tinnitus (2-8). An emerging pattern associated with tinnitus pathology indicates that intense noise exposure leads to cochlear damage and hearing loss, which often is not clinically detected. Decreased cochlear input leads to hyperactive, more responsive central auditory circuits, which is evidenced by functional MRI (fMRI) studies in patients with tinnitus and in vivo recordings in animal models of tinnitus (9-13). Increased spontaneous firing rates, increased evoked responses, and reorganization of tonotopic maps are consistent with decreased inhibition (disinhibition) (1, 14). However, direct evidence that disinhibition mediates these changes is still lacking. In addition, the alternative hypothesis that predicts increased excitation as a potential mechanism in mediating these changes has not been tested (15). Addressing these unexamined questions could lead to pharmacological approaches for treating tinnitus patients.We used flavoprotein autofluorescence (FA) imaging in brain slices prepared from...

Motor cortex is a key brain center involved in motor control in rodents and other mammals, but specific intracortical mechanisms at the microcircuit level are largely unknown. Neuronal expression of hyperpolarization-activated current (I(h)) is cell class specific throughout the nervous system, but in neocortex, where pyramidal neurons are classified in various ways, a systematic pattern of expression has not been identified. We tested whether I(h) is differentially expressed among projection classes of pyramidal neurons in mouse motor cortex. I(h) expression was high in corticospinal neurons and low in corticostriatal and corticocortical neurons, a pattern mirrored by mRNA levels for HCN1 and Trip8b subunits. Optical mapping experiments showed that I(h) attenuated glutamatergic responses evoked across the apical and basal dendritic arbors of corticospinal but not corticostriatal neurons. Due to I(h), corticospinal neurons resonated, with a broad peak at ∼4 Hz, and were selectively modulated by α-adrenergic stimulation. I(h) reduced the summation of short trains of artificial excitatory postsynaptic potentials (EPSPs) injected at the soma, and similar effects were observed for short trains of actual EPSPs evoked from layer 2/3 neurons. I(h) narrowed the coincidence detection window for EPSPs arriving from separate layer 2/3 inputs, indicating that the dampening effect of I(h) extended to spatially disperse inputs. To test the role of corticospinal I(h) in transforming EPSPs into action potentials, we transfected layer 2/3 pyramidal neurons with channelrhodopsin-2 and used rapid photostimulation across multiple sites to synaptically drive spiking activity in postsynaptic neurons. Blocking I(h) increased layer 2/3-driven spiking in corticospinal but not corticostriatal neurons. Our results imply that I(h)-dependent synaptic integration in corticospinal neurons constitutes an intracortical control mechanism, regulating the efficacy with which local activity in motor cortex is transferred to downstream circuits in the spinal cord. We speculate that modulation of I(h) in corticospinal neurons could provide a microcircuit-level mechanism involved in translating action planning into action execution.

SummaryFrontal cortex plays a central role in the control of voluntary movements, which are typically guided by sensory input. Here, we investigate the function of mouse whisker primary motor cortex (wM1), a frontal region defined by dense innervation from whisker primary somatosensory cortex (wS1). Optogenetic stimulation of wM1 evokes rhythmic whisker protraction (whisking), whereas optogenetic inactivation of wM1 suppresses initiation of whisking. Whole-cell membrane potential recordings and silicon probe recordings of action potentials reveal layer-specific neuronal activity in wM1 at movement initiation, and encoding of fast and slow parameters of movements during whisking. Interestingly, optogenetic inactivation of wS1 caused hyperpolarization and reduced firing in wM1, together with reduced whisking. Optogenetic stimulation of wS1 drove activity in wM1 with complex dynamics, as well as evoking long-latency, wM1-dependent whisking. Our results advance understanding of a well-defined frontal region and point to an important role for sensory input in controlling motor cortex.