dii-perfusion-as-method-for-vascular-visualization-ambystoma

DiI Perfusion as a Method for Vascular Visualization in <em>Ambystoma mexicanum</em>

Perfusion techniques have been used for centuries to visualize the circulation of tissues. Axolotl (Ambystoma mexicanum) is a species of salamander that ...

construction-local-field-potential-microelectrodes-for-vivo

Construction of Local Field Potential Microelectrodes for <em>in vivo</em> Recordings from Multiple Brain Structures Simultaneously

Researchers often need to record local field potentials (LFPs) simultaneously from several brain structures. Recording from multiple desired brain regions ...

Harvard Medical School and Brigham and Women's Hospital

University of Virginia

Generalized convulsive status epilepticus is a life-threatening emergency, because recurrent convulsions can cause death or injury. A common form of generalized convulsive status epilepticus is of focal onset. The neuronal circuits activated during seizure spread from the hippocampus, a frequent site of seizure origin, to the bilateral motor cortex, which mediates convulsive seizures, have not been delineated. Status epilepticus was initiated by electrical stimulation of the hippocampus. Neurons transiently activated during seizures were labelled with tdTomato and then imaged following brain slice clearing. Hippocampus was active throughout the episode of status epilepticus. Neuronal activation was observed in hippocampus parahippocampal structures: subiculum, entorhinal cortex and perirhinal cortex, septum, and olfactory system in the initial phase status epilepticus. The tdTomato-labelled neurons occupied larger volumes of the brain as seizures progressed and at the peak of status epilepticus, motor and somatosensory cortex, retrosplenial cortex, and insular cortex also contained tdTomato-labelled neurons. In addition, motor thalamic nuclei such as anterior and ventromedial, midline, reticular, and posterior thalamic nuclei were also activated. Furthermore, circuits proposed to be crucial for systems consolidation of memory: entorhinal cortex, retrosplenial cortex, cingulate gyrus, midline thalamic nuclei and prefrontal cortex were intensely active during periods of generalized tonic-clonic seizures. As the episode of status epilepticus waned, smaller volume of brain was activated. These studies suggested that seizure spread could have occurred via canonical thalamocortical pathway and many cortical structures involved in memory consolidation. These studies may help explain retrograde amnesia following seizures.

Parallel pathways of seizure generalization.

Mapping the circuits underlying the generation and propagation of seizures is critically important for understanding their pathophysiology. We review evidence to suggest that circuits engaged in secondarily generalized seizures are likely to be more complex than those currently proposed. Focal seizures have been proposed to engage canonical thalamocortical circuits that mediate primarily generalized absence seizures, leading to secondarily generalized tonic-clonic seizures. In addition to traveling through the canonical thalamocortical circuits, secondarily generalized seizures could also travel through the striatum, globus pallidus, substantia nigra reticulata, and corpus callosum to the contralateral hemisphere. Recruitment of principal neurons in superficial layers 2/3 of the cortex can play a critical role in corticocortical seizure spread. Understanding the neuronal structures engaged in generating secondarily generalized seizures could provide novel targets for neuromodulation for the treatment of seizures. Furthermore, these sites may be loci of neuronal plasticity facilitating epileptogenesis. This article is part of the Special Issue "Proceedings of the 7th London-Innsbruck Colloquium on Status Epilepticus and Acute Seizures".

Circuits generating secondarily generalized seizures.

There are no detailed descriptions of neuronal circuit active during frontal lobe motor seizures. Using activity reporter mice, local field potential recordings, tissue clearing, viral tracing, and super-resolution microscopy, we found neuronal activation after focal motor to bilateral tonic-clonic seizures in the striatum, globus pallidus externus, subthalamic nucleus, substantia nigra pars reticulata and neurons of the indirect pathway. Seizures preferentially activated dopamine D2 receptor-expressing neurons over D1 in the striatum, which have different projections. Furthermore, the D2 receptor agonist infused into the striatum exerted an anticonvulsant effect. Seizures activate structures via short and long latency loops, and anatomical connections of the seizure focus determine the seizure circuit. These studies, for the first time, show activation of neurons in the striatum, globus pallidus, subthalamic nucleus, and substantia nigra during frontal lobe motor seizures on the cellular level, revealing a complex neuronal activation circuit subject to modulation by the basal ganglia.

Activation of the basal ganglia and indirect pathway neurons during frontal lobe seizures.

There is uncertainty regarding when and which groups of neurons fire synchronously during seizures. While several studies found heterogeneous firing during seizures, others suggested synchronous neuronal firing in the seizure core. We tested whether neuronal activity during seizures is orderly in the direction of the excitatory neuronal connections in the circuit. There are strong excitatory connections laterally within the septotemporally organized lamella and inhibitory trans-lamellar connections in the hippocampus, which allow testing of the connectivity hypothesis. We further tested whether epileptogenesis enhances synchrony and antiseizure drug administration disrupts it. We recorded local field potentials from CA1 pyramidal neurons using a small microelectrode array and kindled rats by a rapid, recurrent hippocampal stimulation protocol. We compared cross-correlation, theta phase synchronization, entropy, and event synchronization. These analyses revealed that the firing pattern was correlated along the lamellar, but not the septotemporal, axis during evoked seizures. During kindling, neuronal synchrony increased along the lamellar axis, while synchrony along the septotemporal axis remained relatively low. Additionally, the theta phase distribution demonstrated that CA1 pyramidal cell firing became preferential for theta oscillation negative peak as kindling progressed in the lamellar direction but not in the trans-lamellar direction. Last, event synchronization demonstrated that neuronal firings along the lamellar axis were more synchronized than those along the septotemporal axis. There was a marked decrease in synchronization and phase preference after treatment with phenytoin and levetiracetam. The synchrony structure of CA1 pyramidal neurons during seizures and epileptogenesis depends on anatomic connectivity and plasticity. We could improve the efficacy of brain stimulation to treat seizures by understanding the structure of synchrony. Electrical stimulation may disrupt seizures by desynchronizing neurons, but there is an uncertainty on which groups of neurons fire synchronously or chaotically during seizures. Here, we demonstrate that neurons linked by excitatory connections fire synchronously during seizures, and this synchrony is modulated by epileptogenesis and antiseizure drugs. Closed-loop brain stimulation carefully targeted to disrupt synchrony may improve the treatment of seizures.

Connectivity and Neuronal Synchrony during Seizures.

Dopamine type 2 receptor (D2R) agonists have anticonvulsant effect, whereas D2R antagonists increase seizure risk, but the mechanism of this action has not been delineated. We tested whether D2R agonists activate parvalbumin (PV)-containing inhibitory interneurons to suppress seizures. We treated frontal lobe onset seizures with a D2R agonist sumanirole, and it suppressed seizures. We used activity reporter TRAP2 mice and found that injection of D2R agonist led to extensive activation of PV interneurons in the cortex and striatum ipsilateral to the seizure focus. D2R agonists activate PV interneurons, which in turn inhibit principal neurons, potentially explaining their anticonvulsant effect.

Anticonvulsant dopamine type 2 receptor agonist activates inhibitory parvalbumin interneurons.

Bilateral synchronous cortical activity occurs during sleep, attention, and seizures. Canonical models place the thalamus at the center of bilateral cortical synchronization because it generates bilateral sleep spindle oscillations and primarily generalized absence seizures. However, classical studies suggest that the corpus callosum mediates bilateral cortical synchronization.

Anastasia Brodovskaya

Department of Orthopedic Surgery

Harvard Medical School and Brigham and Women's Hospital

DiI Perfusion as a Method for Vascular Visualization in Ambystoma mexicanum

Construction of Local Field Potential Microelectrodes for in vivo Recordings from Multiple Brain Structures Simultaneously

Anastasia Brodovskaya

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Orthopedic Surgery

Harvard Medical School and Brigham and Women's Hospital

DiI Perfusion as a Method for Vascular Visualization in Ambystoma mexicanum

Construction of Local Field Potential Microelectrodes for in vivo Recordings from Multiple Brain Structures Simultaneously

Department of Orthopedic Surgery