Michal Harel

Interhemispheric correlations of slow spontaneous neuronal fluctuations revealed in human sensory cortex

Animal studies have shown robust electrophysiological activity in the sensory cortex in the absence of stimuli or tasks. Similarly, recent human functional magnetic resonance imaging (fMRI) revealed widespread, spontaneously emerging cortical fluctuations. However, it is unknown what neuronal dynamics underlie this spontaneous activity in the human brain. Here we studied this issue by combining bilateral single-unit, local field potentials (LFPs) and intracranial electrocorticography (ECoG) recordings in individuals undergoing clinical monitoring. We found slow (<0.1 Hz, following 1/f-like profiles) spontaneous fluctuations of neuronal activity with significant interhemispheric correlations. These fluctuations were evident mainly in neuronal firing rates and in gamma (40–100 Hz) LFP power modulations. Notably, the interhemispheric correlations were enhanced during rapid eye movement and stage 2 sleep. Multiple intracranial ECoG recordings revealed clear selectivity for functional networks in the spontaneous gamma LFP power modulations. Our results point to slow spontaneous modulations in firing rate and gamma LFP as the likely correlates of spontaneous fMRI fluctuations in the human sensory cortex.

Internally generated reactivation of single neurons in human hippocampus during free recall.

The emergence of memory, a trace of things past, into human consciousness is one of the greatest mysteries of the human mind. Whereas the neuronal basis of recognition memory can be probed experimentally in human and nonhuman primates, the study of free recall requires that the mind declare the occurrence of a recalled memory (an event intrinsic to the organism and invisible to an observer). Here, we report the activity of single neurons in the human hippocampus and surrounding areas when subjects first view cinematic episodes consisting of audiovisual sequences and again later when they freely recall these episodes. A subset of these neurons exhibited selective firing, which often persisted throughout and following specific episodes for as long as 12 seconds. Verbal reports of memories of these specific episodes at the time of free recall were preceded by selective reactivation of the same hippocampal and entorhinal cortex neurons. We suggest that this reactivation is an internally generated neuronal correlate for the subjective experience of spontaneous emergence of human recollection.

Large-Scale Mirror-Symmetry Organization of Human Occipito-Temporal Object Areas

We have combined functional maps of retinotopy (eccentricity and meridian mapping), object category, and motion in a group of subjects to explore the large-scale topography of higher-order object areas. Our results reveal seven consistent category-related entities situated in the occipito-temporal cortex adjoining early visual areas. These include two face-related regions, three object-related regions, and two building-related regions. Interestingly, this complex category-related pattern is organized in a large-scale dorso-ventral mirror symmetry of object category. Furthermore, correlating this pattern to the map of visual field eccentricity, we found that the entire network of areas could be related to a single and unified eccentricity map. We hypothesize that this large-scale organization points to a possible development of high-order object areas through extension and specialization of a single proto-representation.

Acetylcholinesterase: From 3D structure to function.

By rapid hydrolysis of the neurotransmitter, acetylcholine, acetylcholinesterase terminates neurotransmission at cholinergic synapses. Acetylcholinesterase is a very fast enzyme, functioning at a rate approaching that of a diffusion-controlled reaction. The powerful toxicity of organophosphate poisons is attributed primarily to their potent inhibition of acetylcholinesterase. Acetylcholinesterase inhibitors are utilized in the treatment of various neurological disorders, and are the principal drugs approved thus far by the FDA for management of Alzheimers disease. Many organophosphates and carbamates serve as potent insecticides, by selectively inhibiting insect acetylcholinesterase. The determination of the crystal structure of Torpedo californica acetylcholinesterase permitted visualization, for the first time, at atomic resolution, of a binding pocket for acetylcholine. It also allowed identification of the active site of acetylcholinesterase, which, unexpectedly, is located at the bottom of a deep gorge lined largely by aromatic residues. The crystal structure of recombinant human acetylcholinesterase in its apo-state is similar in its overall features to that of the Torpedo enzyme; however, the unique crystal packing reveals a novel peptide sequence which blocks access to the active-site gorge.

X‐ray structure of human acid‐β‐glucosidase, the defective enzyme in Gaucher disease

Gaucher disease, the most common lysosomal storage disease, is caused by mutations in the gene that encodes acid‐β‐glucosidase (GlcCerase). Type 1 is characterized by hepatosplenomegaly, and types 2 and 3 by early or chronic onset of severe neurological symptoms. No clear correlation exists between the ∼200 GlcCerase mutations and disease severity, although homozygosity for the common mutations N370S and L444P is associated with non‐ neuronopathic and neuronopathic disease, respectively. We report the X‐ray structure of GlcCerase at 2.0 Å resolution. The catalytic domain consists of a (β/α)8 TIM barrel, as expected for a member of the glucosidase hydrolase A clan. The distance between the catalytic residues E235 and E340 is consistent with a catalytic mechanism of retention. N370 is located on the longest α‐helix (helix 7), which has several other mutations of residues that point into the TIM barrel. Helix 7 is at the interface between the TIM barrel and a separate immunoglobulin‐like domain on which L444 is located, suggesting an important regulatory or structural role for this non‐catalytic domain. The structure provides the possibility of engineering improved GlcCerase for enzyme‐replacement therapy, and for designing structure‐based drugs aimed at restoring the activity of defective GlcCerase.

Neural “Ignition”: Enhanced Activation Linked to Perceptual Awareness in Human Ventral Stream Visual Cortex

Human recognition performance is characterized by abrupt changes in perceptual states. Understanding the neuronal dynamics underlying such transitions could provide important insights into mechanisms of recognition and perceptual awareness. Here we examined patients monitored for clinical purposes with multiple subdural electrodes. The patients participated in a backward masking experiment in which pictures of various object categories were presented briefly followed by a mask. We recorded ECoG from 445 electrodes placed in 11 patients. We found a striking increase in gamma power (30-70 Hz) and evoked responses specifically associated with successful recognition. The enhanced activation occurred 150-200 ms after stimulus onset and consistently outlasted the stimulus presentation. We propose that the gamma and evoked potential activations reflect a rapid increase in recurrent neuronal activity that plays a critical role in the emergence of a recognizable visual percept in conscious awareness.

The binding site of acetylcholine receptor as visualized in the X-Ray structure of a complex between alpha-bungarotoxin and a mimotope peptide.

We have determined the crystal structure at 1.8 A resolution of a complex of alpha-bungarotoxin with a high affinity 13-residue peptide that is homologous to the binding region of the alpha subunit of acetylcholine receptor. The peptide fits snugly to the toxin and adopts a beta hairpin conformation. The structures of the bound peptide and the homologous loop of acetylcholine binding protein, a soluble analog of the extracellular domain of acetylcholine receptor, are remarkably similar. Their superposition indicates that the toxin wraps around the receptor binding site loop, and in addition, binds tightly at the interface of two of the receptor subunits where it inserts a finger into the ligand binding site, thus blocking access to the acetylcholine binding site and explaining its strong antagonistic activity.

Crystal structure of thioflavin T bound to the peripheral site of Torpedo californica acetylcholinesterase reveals how thioflavin T acts as a sensitive fluorescent reporter of ligand binding to the acylation site.

Acetylcholinesterase plays a key role in cholinergic synaptic transmission by hydrolyzing the neurotransmitter acetylcholine with one of the highest known catalytic rate constants. Hydrolysis occurs in a narrow and deep gorge that contains two sites of ligand binding: A peripheral site, or P-site, near the gorge entrance that contributes to catalytic efficiency both by transiently trapping substrate molecules as they enter the gorge and by allosterically accelerating the transfer of the substrate acyl group to a serine hydroxyl in an acylation site or A-site at the base of the gorge. Thioflavin T is a useful reporter of ligand interactions with the A-site. It binds specifically to the P-site with fluorescence that is enhanced approximately 1000-fold over that of unbound thioflavin T, and the enhanced fluorescence is quenched 1.5- to 4-fold when another ligand binds to the A-site in a ternary complex. To clarify the structural basis of this advantageous signal change, we here report the X-ray structure of the complex of thioflavin T with Torpedo californica acetylcholinesterase. The two aromatic rings in thioflavin T are coplanar and are packed snugly parallel to the aromatic side chains of Trp279, Tyr334, and Phe330. Overlays of this structure with the crystal structures of Torpedo californica acetylcholinesterase complexes with either edrophonium or m-( N, N, N-trimethylammonio)-2,2,2-trifluoroacetophenone, two small aromatic ligands that bind specifically to the A-site, indicate that the phenyl side chain of Phe330 must rotate to sterically accommodate both thioflavin T and the A-site ligand in the ternary complex. This rotation may allow some relaxation of the strict coplanarity of the aromatic rings in the bound thioflavin T and result in partial quenching of its fluorescence.

Retinotopic Axis Specificity and Selective Clustering of Feedback Projections from V2 to V1 in the Owl Monkey

Cortical maps and feedback connections are ubiquitous features of the visual cerebral cortex. The role of the feedback connections, however, is unclear. This study was aimed at revealing possible organizational relationships between the feedback projections from area V2 and the functional maps of orientation and retinotopy in area V1. Optical imaging of intrinsic signals was combined with cytochrome oxidase histochemistry and connectional anatomy in owl monkeys. Tracer injections were administered at orientation-selective domains in regions of pale and thick cytochrome oxidase stripes adjacent to the border between these stripes. The feedback projections from V2 were found to be more diffuse than the intrinsic horizontal connections within V1, but they nevertheless demonstrated clustering. The clusters of feedback axons projected preferentially to interblob cytochrome oxidase regions. The distribution of preferred orientations of the recipient domains in V1 was broad but appeared biased toward values similar to the preferred orientation of the projecting cells in V2. The global spatial distribution of the feedback projections in V1 was anisotropic. The major axis of anisotropy was systematically parallel to a retinotopic axis in V1 corresponding to the preferred orientation of the cells of origin in V2. We conclude that the feedback connections from V2 to V1 might play a role in enhancing the response in V1 to collinear contour elements.

The synaptic acetylcholinesterase tetramer assembles around a polyproline II helix

Functional localization of acetylcholinesterase (AChE) in vertebrate muscle and brain depends on interaction of the tryptophan amphiphilic tetramerization (WAT) sequence, at the C‐terminus of its major splice variant (T), with a proline‐rich attachment domain (PRAD), of the anchoring proteins, collagenous (ColQ) and proline‐rich membrane anchor. The crystal structure of the WAT/PRAD complex reveals a novel supercoil structure in which four parallel WAT chains form a left‐handed superhelix around an antiparallel left‐handed PRAD helix resembling polyproline II. The WAT coiled coils possess a WWW motif making repetitive hydrophobic stacking and hydrogen‐bond interactions with the PRAD. The WAT chains are related by an ∼4‐fold screw axis around the PRAD. Each WAT makes similar but unique interactions, consistent with an asymmetric pattern of disulfide linkages between the AChE tetramer subunits and ColQ. The P59Q mutation in ColQ, which causes congenital endplate AChE deficiency, and is located within the PRAD, disrupts crucial WAT–WAT and WAT–PRAD interactions. A model is proposed for the synaptic AChET tetramer.