Claustrum projections to the anterior cingulate modulate nociceptive and pain-associated behavior.

The anterior cingulate cortex (ACC) is critical for the perception and unpleasantness of pain.1,2,3,4,5,6 It receives nociceptive information from regions such as the thalamus and amygdala and projects to several cortical and subcortical regions of the pain neuromatrix.7,8 ACC hyperexcitability is one of many functional changes associated with chronic pain, and experimental activation of ACC pyramidal cells produces hypersensitivity to innocuous stimuli (i.e., allodynia).9,10,11,12,13,14 A less-well-studied projection to the ACC arises from a small forebrain region, the claustrum.15,16,17,18,19,20 Stimulation of excitatory claustrum projection neurons preferentially activates GABAergic interneurons, generating feed-forward inhibition onto excitatory cortical networks.21,22,23,24 Previous work has shown that claustrocingulate projections display altered activity in prolonged pain25,26,27; however, it remains unclear whether and how the claustrum participates in nociceptive processing and high-order pain behaviors. Inhibition of ACC activity reverses mechanical allodynia in animal models of persistent and neuropathic pain,1,9,28 suggesting claustrum inputs may function to attenuate pain processing. In this study, we sought to define claustrum function in acute and chronic pain. We found enhanced claustrum activity after a painful stimulus that was attenuated in chronic inflammatory pain. Selective inhibition of claustrocingulate projection neurons enhanced acute nociception but blocked pain learning. Inversely, chemogenetic activation of claustrocingulate neurons had no effect on basal nociception but rescued inflammation-induced mechanical allodynia. Together, these results suggest that claustrocingulate neurons are a critical component of the pain neuromatrix, and dysregulation of this connection may contribute to chronic pain.

Transplantation of glutamatergic neuronal precursor cells in the paraventricular thalamus and claustrum facilitates awakening with recovery of consciousness.

BACKGROUND: Stem cells offer a promising therapeutic strategy for patients with disorders of consciousness (DOC) after severe traumatic brain injury (TBI), but the optimal transplantation sites and cells are not clear. Although the paraventricular thalamus (PVT) and claustrum (CLA) are associated with consciousness and are candidate transplantation targets, few studies have been designed to investigate this possibility. METHODS: Controlled cortical injury (CCI) was performed to establish a mouse model of DOC. CCI-DOC paradigm was established to investigate the role of excitatory neurons of PVT and CLA in disorders of consciousness. The role of excitatory neuron transplantation in promoting arousal and recovery of consciousness was determined by optogenetics, chemogenetics, electrophysiology, Western blot, RT-PCR, double immunofluorescence labeling, and neurobehavioral experiments. RESULTS: After CCI-DOC, neuronal apoptosis was found to be concentrated in the PVT and CLA. Prolonged awaking latency and cognitive decline were also seen after destruction of the PVT and CLA, suggesting that the PVT and CLA may be key nuclei in DOC. Awaking latency and cognitive performance could be altered by inhibiting or activating excitatory neurons, implying that excitatory neurons may play an important role in DOC. Furthermore, we found that the PVT and CLA function differently, with the PVT mainly involved in arousal maintenance while the CLA plays a role mainly in the generation of conscious content. Finally, we found that by transplanting excitatory neuron precursor cells in the PVT and CLA, respectively, we could facilitate awakening with recovery of consciousness, which was mainly manifested by shortened awaking latency, reduced duration of loss of consciousness (LOC), enhanced cognitive ability, enhanced memory, and improved limb sensation. CONCLUSION: In this study, we found that the deterioration in the level and content of consciousness after TBI was associated with a large reduction in glutamatergic neurons within the PVT and CLA. Transplantation of glutamatergic neuronal precursor cells could play a beneficial role in promoting arousal and recovery of consciousness. Thus, these findings have the potential to provide a favorable basis for promoting awakening and recovery in patients with DOC.

Preferential arborization of dendrites and axons of parvalbumin- and somatostatin-positive GABAergic neurons within subregions of the mouse claustrum.

The claustrum coordinates the activities of individual cortical areas through abundant reciprocal connections with the cerebral cortex. Although these excitatory connections have been extensively investigated in three subregions of the claustrum-core region and dorsal and ventral shell regions-the contribution of GABAergic neurons to the circuitry in each subregion remains unclear. Here, we examined the distribution of GABAergic neurons and their dendritic and axonal arborizations in each subregion. Combining in situ hybridization with immunofluorescence histochemistry showed that approximately 10% of neuronal nuclei-positive cells expressed glutamic acid decarboxylase 67 mRNA across the claustral subregions. Approximately 20%, 30%, and 10% of GABAergic neurons were immunoreactive for parvalbumin (PV), somatostatin (SOM), and vasoactive intestinal polypeptide, respectively, in each subregion, and these neurochemical markers showed little overlap with each other. We then reconstructed PV and SOM neurons labeled with adeno-associated virus vectors. The dendrites and axons of PV and SOM neurons were preferentially localized to their respective subregions where their cell bodies were located. Furthermore, the axons were preferentially extended in a rostrocaudal direction, whereas the dendrites were relatively isotropic. The present findings suggest that claustral PV and SOM neurons might execute information processing separately within the core and shell regions.

Mice Lacking Connective Tissue Growth Factor in the Forebrain Exhibit Delayed Seizure Response, Reduced C-Fos Expression and Different Microglial Phenotype Following Acute PTZ Injection.

Connective tissue growth factor (CTGF) plays important roles in the development and regeneration of the connective tissue, yet its function in the nervous system is still not clear. CTGF is expressed in some distinct regions of the brain, including the dorsal endopiriform nucleus (DEPN) which has been recognized as an epileptogenic zone. We generated a forebrain-specific Ctgf knockout (FbCtgf KO) mouse line in which the expression of Ctgf in the DEPN is eliminated. In this study, we adopted a pentylenetetrazole (PTZ)-induced seizure model and found similar severity and latencies to death between FbCtgf KO and WT mice. Interestingly, there was a delay in the seizure reactions in the mutant mice. We further observed reduced c-fos expression subsequent to PTZ treatment in the KO mice, especially in the hippocampus. While the densities of astrocytes and microglia in the hippocampus were kept constant after acute PTZ treatment, microglial morphology was different between genotypes. Our present study demonstrated that in the FbCtgf KO mice, PTZ failed to increase neuronal activity and microglial response in the hippocampus. Our results suggested that inhibition of Ctgf function may have a therapeutic potential in preventing the pathophysiology of epilepsy.

Potential Role of the Amygdala and Posterior Claustrum in Exercise Intensity-dependent Cardiovascular Regulation in Rats.

Tuning of the cardiovascular response is crucial to maintain performance during high-intensity exercise. It is well known that the nucleus of the solitary tract (NTS) in the brainstem medulla plays a central role in cardiovascular regulation; however, where and how upper brain regions form circuits with NTS and coordinately control cardiovascular responses during high-intensity exercise remain unclear. Here focusing on the amygdala and claustrum, we investigated part of the mechanism for regulation of the cardiovascular system during exercise. In rats, c-Fos immunostaining was used to examine whether the amygdala and claustrum were activated during treadmill exercise. Further, we examined arterial pressure responses to electrical and chemical stimulation of the claustrum region. We also confirmed the anatomical connections between the amygdala, claustrum, and NTS by retrograde tracer injections. Finally, we performed simultaneous electrical stimulation of the claustrum and amygdala to examine their functional connectivity. c-Fos expression was observed in the amygdala and the posterior part of the claustrum (pCL), but not in the anterior part, in an exercise intensity-dependent manner. pCL stimulation induced a depressor response. Using a retrograde tracer, we confirmed direct projections from the amygdala to the pCL and NTS. Simultaneous stimulation of the central nucleus of the amygdala and pCL showed a greater pressor response compared with the stimulation of the amygdala alone. These results suggest the amygdala and pCL are involved in different phases of exercise. More speculatively, these areas might coordinately tune cardiovascular responses that help maintain performance during high-intensity exercise.

Microneuroanatomy of the Anterior Frontal Laser Trajectory to the Insula.

BACKGROUND: Magnetic resonance imaging-guided laser interstitial thermal therapy (LITT) is an emerging minimally invasive procedure for the treatment of deep intracranial lesions. Insular lesions are challenging to treat because of the risk of damaging important surrounding structures. The precise knowledge of the neural structures that are at risk along the trajectory and during the ablation is essential to reduce associated complications. This study aims to describe the relevant anatomy of the anterior frontal LITT trajectory to the insular region by using sectional anatomy and fiber dissection technique. METHODS: Three silicone-injected cadaveric heads were used to implant laser catheters bilaterally to the insular region by using a frameless stereotactic technique from a frontal approach. Sections were cut in both the oblique axial plane parallel to the trajectory and in the coronal plane. White matter fiber dissections were used to establish the tracts related to the laser trajectory from lateral to medial and medial to lateral. RESULTS: Supraorbital regions were selected as entry points. After crossing the frontal bone, the track intersected the inferior frontal lobe. The catheter was illustrated reaching the insular region medial to the inferior fronto-occipital fasciculus and insular cortex, and superior to the uncinate fasciculus. The uncinate fasciculus, extreme capsule, claustrum, external capsule, and putamen were traversed, preserving the major vascular structures. CONCLUSIONS: Independent of the insular area treated, an understanding of the neuroanatomy related to the anterior frontal laser trajectory is essential to improve the ability to perform LITT of this challenging region.

Input-output organization of the mouse claustrum.

Progress in determining the precise organization and function of the claustrum (CLA) has been hindered by the difficulty in reliably targeting these neurons. To overcome this, we used a projection-based targeting strategy to selectively label CLA principal neurons. Combined with adeno-associated virus (AAV) and monosynaptic rabies tracing techniques, we systematically examined the pre-synaptic input and axonal output of this structure. We found that CLA neurons projecting to retrosplenial cortex (RSP) collateralize extensively to innervate a variety of higher-order cortical regions. No subcortical labeling was found, with the exception of sparse terminals in the basolateral amygdala (BLA). This pattern of output was similar to cingulate- and visual cortex-projecting CLA neurons, suggesting a common targeting scheme among these projection-defined populations. Rabies virus tracing directly demonstrated widespread synaptic inputs to RSP-projecting CLA neurons from both cortical and subcortical areas. The strongest inputs arose from classically defined limbic regions, including medial prefrontal cortex, anterior cingulate, BLA, ventral hippocampus, and neuromodulatory systems such as the dorsal raphe and cholinergic basal forebrain. These results suggest that the CLA may integrate information related to the emotional salience of stimuli and may globally modulate cortical state by broadcasting its output uniformly across a variety of higher cognitive centers.