Lumbar V3 interneurons provide direct excitatory synaptic input onto thoracic sympathetic preganglionic neurons, linking locomotor, and autonomic spinal systems.

Although sympathetic autonomic systems are activated in parallel with locomotion, the neural mechanisms mediating this coordination are incompletely understood. Sympathetic preganglionic neurons (SPNs), primarily located in the intermediate laminae of thoracic and upper lumbar segments (T1-L2), increase activation of tissues and organs that provide homeostatic and metabolic support during movement and exercise. Recent evidence suggests integration between locomotor and autonomic nuclei occurs within the brainstem, initiating both descending locomotor and sympathetic activation commands. However, both locomotor and sympathetic autonomic spinal systems can be activated independent of supraspinal input, in part due to a distributed network involving propriospinal neurons. Whether an intraspinal mechanism exists to coordinate activation of these systems is unknown. We hypothesized that ascending spinal neurons located in the lumbar region provide synaptic input to thoracic SPNs. Here, we demonstrate that synaptic contacts from locomotor-related V3 interneurons (INs) are present in all thoracic laminae. Injection of an anterograde tracer into lumbar segments demonstrated that 8-20% of glutamatergic input onto SPNs originated from lumbar V3 INs and displayed a somatotopographical organization of synaptic input. Whole cell patch clamp recording in SPNs demonstrated prolonged depolarizations or action potentials in response to optical activation of either lumbar V3 INs in spinal cord preparations or in response to optical activation of V3 terminals in thoracic slice preparations. This work demonstrates a direct intraspinal connection between lumbar locomotor and thoracic sympathetic networks and suggests communication between motor and autonomic systems may be a general function of the spinal cord.

Intrinsic brainstem circuits comprised of Chx10-expressing neurons contribute to reticulospinal output in mice.

Glutamatergic reticulospinal neurons in the gigantocellular reticular nucleus (GRN) of the medullary reticular formation can function as command neurons, transmitting motor commands to spinal cord circuits to instruct movement. Recent advances in our understanding of this neuron-dense region have been facilitated by the discovery of expression of the transcriptional regulator, Chx10, in excitatory reticulospinal neurons. Here, we address the capacity of local circuitry in the GRN to contribute to reticulospinal output. We define two subpopulations of Chx10-expressing neurons in this region, based on distinct electrophysiological properties and soma size (small and large), and show that these populations correspond to local interneurons and reticulospinal neurons, respectively. Using focal release of caged glutamate combined with patch clamp recordings, we demonstrated that Chx10 neurons form microcircuits in which the Chx10 local interneurons project to and facilitate the firing of Chx10 reticulospinal neurons. We discuss the implications of these microcircuits in terms of movement selection.NEW & NOTEWORTHY Reticulospinal neurons in the medullary reticular formation integrate inputs from higher regions to effectively instruct spinal motor circuits. Using photoactivation of neurons in brainstem slices, we studied connectivity of reticular formation neurons that express the transcriptional regulator, Chx10. We show that a subpopulation of these neurons functions as local interneurons that affect descending commands. The results shed light on the internal organization and microcircuit formation of reticular formation neurons.

Schizophrenia-associated LRRTM1 regulates cognitive behavior through controlling synaptic function in the mediodorsal thalamus.

Reduced activity of the mediodorsal thalamus (MD) and abnormal functional connectivity of the MD with the prefrontal cortex (PFC) cause cognitive deficits in schizophrenia. However, the molecular basis of MD hypofunction in schizophrenia is not known. Here, we identified leucine-rich-repeat transmembrane neuronal protein 1 (LRRTM1), a postsynaptic cell-adhesion molecule, as a key regulator of excitatory synaptic function and excitation-inhibition balance in the MD. LRRTM1 is strongly associated with schizophrenia and is highly expressed in the thalamus. Conditional deletion of Lrrtm1 in the MD in adult mice reduced excitatory synaptic function and caused a parallel reduction in the afferent synaptic activity of the PFC, which was reversed by the reintroduction of LRRTM1 in the MD. Our results indicate that chronic reduction of synaptic strength in the MD by targeted deletion of Lrrtm1 functionally disengages the MD from the PFC and may account for cognitive, social, and sensorimotor gating deficits, reminiscent of schizophrenia.

Multistable properties of human subthalamic nucleus neurons in Parkinson's disease.

To understand the function and dysfunction of neural circuits, it is necessary to understand the properties of the neurons participating in the behavior, the connectivity between these neurons, and the neuromodulatory status of the circuits at the time they are producing the behavior. Such knowledge of human neural circuits is difficult, at best, to obtain. Here, we study firing properties of human subthalamic neurons, using microelectrode recordings and microstimulation during awake surgery for Parkinson's disease. We demonstrate that low-amplitude, brief trains of microstimulation can lead to persistent changes in neuronal firing behavior including switching between firing rates, entering silent periods, or firing several bursts then entering a silent period. We suggest that these multistable states reflect properties of finite state machines and could have implications for the function of circuits involving the subthalamic nucleus. Furthermore, understanding these states could lead to therapeutic strategies aimed at regulating the transitions between states.

Sub-populations of Spinal V3 Interneurons Form Focal Modules of Layered Pre-motor Microcircuits.

Layering of neural circuits facilitates the separation of neurons with high spatial sensitivity from those that play integrative temporal roles. Although anatomical layers are readily identifiable in the brain, layering is not structurally obvious in the spinal cord. But computational studies of motor behaviors have led to the concept of layered processing in the spinal cord. It has been postulated that spinal V3 interneurons (INs) play multiple roles in locomotion, leading us to investigate whether they form layered microcircuits. Using patch-clamp recordings in combination with holographic glutamate uncaging, we demonstrate focal, layered modules, in which ventromedial V3 INs form synapses with one another and with ventrolateral V3 INs, which in turn form synapses with ipsilateral motoneurons. Motoneurons, in turn, provide recurrent excitatory, glutamatergic input to V3 INs. Thus, ventral V3 interneurons form layered microcircuits that could function to ensure well-timed, spatially specific movements.

Reticulospinal Systems for Tuning Motor Commands.

The pontomedullary reticular formation (RF) is a key site responsible for integrating descending instructions to execute particular movements. The indiscrete nature of this region has led not only to some inconsistencies in nomenclature, but also to difficulties in understanding its role in the control of movement. In this review article, we first discuss nomenclature of the RF, and then examine the reticulospinal motor command system through evolution. These command neurons have direct monosynaptic connections with spinal interneurons and motoneurons. We next review their roles in postural adjustments, walking and sleep atonia, discussing their roles in movement activation or inhibition. We propose that knowledge of the internal organization of the RF is necessary to understand how the nervous system tunes motor commands, and that this knowledge will underlie strategies for motor functional recovery following neurological injuries or diseases.

Case Studies in Neuroscience: Evidence of motor thalamus reorganization following bilateral forearm amputations.

Following injury, functional improvement can result from central nervous system plasticity. Use-dependent plasticity of motor systems is evident, for example, in recovery of function resulting from rehabilitative interventions. Here, we present a single patient who underwent bilateral microelectrode-guided stereotactic implantation of deep brain stimulating leads for the treatment of essential tremor 52 yr following bilateral arm amputations. The tremor affected his upper extremities and had rendered him unable to perform fine motor tasks with his prostheses, significantly reducing his independence. We found a large territory of neurons in the ventral intermediate nucleus of his thalamus that responded to shoulder protraction, the movement that he used to control fine motor movements of his terminal hook prostheses. We propose that reorganization of this motor nucleus may have occurred secondary to a use-dependent gain of function in neurons that were previously involved in hand movement. NEW & NOTEWORTHY We had a unique opportunity to record neurons in the ventrointermediate (Vim) motor nucleus of thalamus in a patient with essential tremor, decades following bilateral forearm amputations. We demonstrate that a large region of Vim is active during shoulder protraction-the movement used to operate the patient's mechanical prostheses. We suggest that this provides evidence of human motor thalamic plasticity.

Altered transcription of glutamatergic and glycinergic receptors in spinal cord dorsal horn following spinal cord transection is minimally affected by passive exercise of the hindlimbs.

Gene expression is altered following a spinal transection (STx) in both motor and sensory systems. Exercise has been shown to influence gene expression in both systems post-STx. Gene expression alterations have also been shown in the dorsal root ganglia and nociceptive laminae of the spinal cord following either an incomplete spinal cord injury (SCI) or a contusive SCI. However, the effect of STx and exercise on gene expression in spinal cord laminae I-III has not fully been examined. Therefore, the purpose of this study was to determine whether gene expression in laminae I-III is altered following STx and determine whether superimposed passive exercise of the hindlimbs would influence gene expression post-STx in laminae I-III. Laser capture microdissection was used to selectively harvest laminae I-III of lumbar spinal cord sections, and quantitative RT-PCR was used to examine relative expression of 23 selected genes in samples collected from control, STx and STx plus exercise rats. We demonstrate that post-STx, gene expression for metabotropic glutamate receptors 1, 5 and 8 were up-regulated, whereas ionotropic glutamatergic receptor (Glur2) and glycinergic subunit GLRA1 expression was down-regulated. Daily exercise attenuated the down-regulation of Glur2 gene expression in laminae I-III. Our results demonstrate that in a STx model, gene expression is altered in laminae I-III and that although passive exercise influences gene expression in both the motor and sensory systems, it had a minimal effect on gene expression in laminae I-III post-STx.

α-Motoneurons maintain biophysical heterogeneity in obesity and diabetes in Zucker rats.

Small-diameter sensory dysfunction resulting from diabetes has received much attention in the literature, whereas the impact of diabetes on α-motoneurons (MN) has not. In addition, the chance of developing insulin resistance and diabetes is increased in obesity. No study has examined the impact of obesity or diabetes on the biophysical properties of MN. Lean Zucker rats and Zucker diabetic fatty (ZDF) rats were separated into lean, obese (ZDF fed standard chow), and diabetic (ZDF fed high-fat diet that led to diabetes) groups. Glass micropipettes recorded hindlimb MN properties from identified flexor and extensor MN. MN were separated within their groups on the basis of input conductance, which created high- and low-input conductance subpopulations for each. A significant shorter (20%) afterhyperpolarization half-decay (AHP1/2) was found in low-conductance MN for the diabetic group only, whereas AHP½ tended to be shorter in the obese group (19%). Significant positive correlations were found among rheobase and input conductance for both lean and obese animals. No differences were found between the groups for afterhyperpolarization amplitude (AHPamp), input conductance, rheobase, or any of the rhythmic firing properties (frequency-current slope and spike-frequency adaptation index). MN properties continue to be heterogeneous in obese and diabetic animals. Obesity does not seem to influence lumbar MN. Despite the resistance of MN to the impact of diabetes, the reduced AHP1/2 decay and the tendency for a reduction in AHPamp may be the first sign of change to MN function.NEW & NOTEWORTHY Knowledge about the impact of obesity and diabetes on the biophysical properties of motoneurons is lacking. We found that diabetes reduces the duration of the afterhyperpolarization and that motoneuron function is unchanged by obesity. A reduced afterhyperpolarization may impact discharge characteristics and may be the first sign of change to motoneuron function.

Neurochemical excitation of thoracic propriospinal neurons improves hindlimb stepping in adult rats with spinal cord lesions.

Using an in vitro neonatal rat brainstem-spinal cord preparation, we previously showed that cervicothoracic propriospinal neurons contribute to descending transmission of the bulbospinal locomotor command signal, and neurochemical excitation of these neurons facilitates signal propagation. The present study examined the relevance of these observations to adult rats in vivo. The first aim was to determine the extent to which rats are able to spontaneously recover hindlimb locomotor function in the presence of staggered contralateral hemisections (left T2-4 and right T9-11) designed to abolish all long direct bulbospinal projections. The second aim was to determine whether neurochemical excitation of thoracic propriospinal neurons in such animals facilitates hindlimb stepping. In the absence of intrathecal drug injection, all animals (n=24) displayed some degree of hindlimb recovery ranging from weak ankle movements to brief periods of unsupported hindlimb stepping on the treadmill. The effect of boluses of neurochemicals delivered via an intrathecal catheter (tip placed midway between the rostral and caudal thoracic hemisections) was examined at post-lesion weeks 3, 6 and 9. Quipazine was particularly effective facilitating hindlimb stepping. Subsequent complete transection above the rostral (n=3) or caudal (n=2) hemisections at week 9 had no consistent effect on drug-free locomotor performance, but the facilitatory effect of drug injection decreased in 4/5 animals. Two animals underwent complete transection at T3 as the first and only surgery and implantation of two intrathecal catheters targeted to the mid-thoracic and lumbar regions, respectively. A similar facilitatory effect on stepping was observed in response to drugs administered via either catheter. The results indicate that partial spontaneous recovery of stepping occurs in adult rats after abolishing all long direct bulbospinal connections, in contrast to previous studies suggesting that hindlimb stepping after dual hemisections either does not occur or is observed only if the second hemisection surgery is delayed relative to the first. The results support the hypothesis that artificial modulation of propriospinal neuron excitability may facilitate recovery of motor function after spinal cord injury. However, whether this facilitation is due to enhanced transmission of a descending locomotor signal or is the result of excitation of thoracolumbar circuits independent of supraspinal influence, requires further study.

Serotonin receptor and KCC2 gene expression in lumbar flexor and extensor motoneurons posttransection with and without passive cycling.

Sacrocaudal motoneuron gene expression is altered following a spinal transection. Of interest here is the regulation of serotonin (5-HT) receptors (R), glutamate receptor, metabotropic 1 (mGluR1), and potassium-chloride cotransporter (KCC2), which mediate motoneuron excitability, locomotor recovery, and spasticity posttransection. The examination of these genes in lumbar motoneurons posttransection has not been studied, which is necessary for developing potential pharmacological interventions aimed at restoring locomotion and/or reducing spasticity. Also, if activity is to be used to promote recovery or reduce spasticity postinjury, a further examination of neuromuscular activity on gene expression posttransection is warranted. The purpose of this study was to examine motoneuronal gene expression of 5-HT receptors, KCC2, and mGluR1 at 3 mo following a complete thoracic spinal cord transection, with and without the inclusion of daily passive cycling. Physiological hindlimb extensor and flexor motoneurons were differentially identified with two retrograde fluorescent tracers, allowing for the identification and separate harvesting of extensor and flexor motoneurons with laser capture microdissection and the subsequent examination of mRNA content using quantitative RT-PCR analysis. We demonstrate that posttransection 5-HT1AR, 5-HT2CR, and mGluR1 expression was downregulated, whereas the 5-HT2AR was upregulated. These alterations in gene expression were observed in both flexor and extensor motoneurons, whereas passive cycling influenced gene expression in extensor but not flexor motoneurons. Passive cycling in extensor motoneurons further enhanced 5-HT2AR expression and increased 5-HT7R and KCC2 expression. Our results demonstrate that passive cycling influences serotonin receptor and KCC2 gene expression and that extensor motoneurons compared with flexor motoneurons may be more plastic to activity-based interventions posttransection.

Daily passive cycling attenuates the hyperexcitability and restores the responsiveness of the extensor monosynaptic reflex to quipazine in the chronic spinally transected rat.

Activity-based interventions such as locomotor training or passive cycling have a positive influence on the spinal circuitry and recovery following a spinal cord injury (SCI). The use of quipazine in combination with exercise training has demonstrated a greater functional recovery than has exercise training alone. However, the influence of exercise or training on the responsiveness of the spinal cord to quipazine has not been examined following a chronic spinal transection. The purpose of this study was to characterize the flexor and extensor monosynaptic reflex (MSR) response pre- and post-quipazine in chronic complete spinally transected rats that either underwent daily passive cycling for 3 months or did not receive passive cycling. Following a chronic spinal transection, the extensor MSR demonstrated a hyperreflexive response (fivefold increase) to afferent stimuli, and did not respond to quipazine injection. With daily passive cycling, the extensor MSR hyperexcitability was attenuated, and the MSR amplitude increased 72% following quipazine injection (p<0.004), which was comparable to the extensor MSR response (94%) in the control group. For both chronic spinal transection groups, the flexor MSR amplitudes were not altered following quipazine injection, whereas in the control group the flexor MSR amplitude increased 86% in response to quipazine (p<0.004). These results demonstrate that passive cycling attenuates the hyperreflexive response of the extensor MSR following a chronic SCI, and restores the MSR response to quipazine.

Removal of supraspinal input reveals a difference in the flexor and extensor monosynaptic reflex response to quipazine independent of motoneuron excitation.

The purpose of this study was to determine if quipazine, a serotonergic agonist, differentially modulates flexor and extensor motor output. This was achieved by examining the monosynaptic reflex (MSR) of the tibial (extensor) and peroneal (flexor) nerves, by determining the basic and rhythmic properties of extensor and flexor motoneurons, and by recording extracellular Ia field potentials of the tibial and peroneal nerves in the in vivo adult decerebrate rat in both spinal intact and acute spinalized preparations. In the spinal intact preparation, the tibial and peroneal MSR amplitude significantly increased compared with baseline in response to quipazine, with no difference between nerves (P < 0.05). In the spinalized preparation, the MSR was significantly increased in both the tibial and peroneal nerves with the latter increasing more than the former (5.7 vs. 3.6 times; P < 0.05). Intracellular motoneuron experiments demonstrated that rheobase decreased, while input resistance, afterhyperpolarization amplitude, and the firing rate at a given current injection increased in motoneurons following quipazine administration with no differences between extensor and flexor motoneurons. Both the tibial and peroneal nerve extracellular Ia field potentials increased with the peroneal demonstrating a significantly greater increase (7 vs. 38%; P < 0.05) following quipazine. It is concluded that in the spinal intact preparation quipazine does not have a differential effect on flexor or extensor motor output. However, in the acute spinalized preparation, quipazine preferentially affects the flexor MSR compared with the extensor MSR, likely due to the removal of a descending tonic inhibition on flexor Ia afferents.

Life-long caloric restriction: Effect on age-related changes in motoneuron numbers, sizes and apoptotic markers.

The first purpose of this study was to determine the effect of advanced age (31 months) on the number of motoneurons in the lumbar enlargement of the rat and to determine if motoneurons die via apoptosis with age. The second purpose was to determine if caloric restriction (CR) would attenuate any observed age-related changes in motoneuron numbers or markers of apoptosis and ROS damage. Using immunohistochemistry to identify choline acetyltransferase (ChAT) - positive motoneurons in the ventro-lateral horn larger than 15µm in diameter and having a clear soma and nucleus were sized and counted. Western blots were used to quantify markers of ROS, apoptosis and autophagy in the ventral horn of the lumbar enlargement. The results suggest that the total number of motoneurons in the rat lumbar enlargement does not significantly decrease with age. Also at the time of sacrifice, aged motoneurons were actively undergoing apoptosis through the intrinsic pathway, in a caspase-dependent manner. CR was able to attenuate the increase in body weight, body weight/muscle mass ratio and the level of activate caspase-3 associated with age. CR also reduced the level of heat shock protein 27, oxoguanine glycosylase 1, cytochrome c and LC3B-I.