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Impact of identified interneurons on cellular network mechanisms in the human and rodent neocortex

Final Report Summary - INTERIMPACT (Impact of identified interneurons on cellular network mechanisms in the human and rodent neocortex)

The group initially focused on genetic fingerprinting of electrophysiologically and anatomically identified neurogliaform cells in the rat cerebral cortex. They developed a method capable of determining the full molecular portrait of neurogliaform cells with oligocellular gene-chip technology and validated a number of previously known marker genes with the further development of single cell based digital PCR technology capable of determining mRNA or miRNA expression in single neurons after patch-clamp recording at a single molecule level. Importantly, this effort resulted in finding the activity of genes previously unassociated with the function of GABAergic neurons. The first example of these genes is the ins2 gene, the only gene capable of producing insulin in the rat and thus it is equivalent to the ins gene in human. The group found that insulin is strongly expressed in GABAergic neurogliaform cells detected by single cell digital polymerase chain reaction and immunocytochemistry. The insulin release enhancer antidiabetic drug glibenclamide promotes de novo insulin synthesis in brain slices and glibenclamide application to neurogliaform cells mimicked the excitation suppressing effect of external insulin on local microcircuits via insulin receptors. Identification of potential local sources of insulin in the cerebral cortex appears a promising avenue which the group plans to follow in the coming year to be completed by determining further mechanisms of locally released insulin action in the microcircuit.
A major effort of the group’s activities in the second half of the grant period focused on unique aspects of the human cerebral cortex. Classic theories link cognitive abilities to synaptic properties and human-specific biophysical features of synapses might contribute to the unparalleled performance of the human cerebral cortex. Paired recordings between human excitatory neurons and their postsynaptic target cells and multiple probability fluctuation analysis revealed that the quantal size in synaptic communication were similar. However, human pyramidal cell to fast-spiking interneuron synapses had 4-times more functional release sites in human connections compared to rats. These connections were mediated on average by three synaptic contacts in both species. Each presynaptic active zone contained 6.2 release sites in human, but only 1.6 in rats. Ultrastuctural analysis of serial sections and electron microscopic tomography showed that an active zone harbors 4 docked vesicles in human, but only a single one in rats. Consequently, a single functional release site as defined by Bernard Katz occupies ~0.012 μm2 in the human presynaptic active zone and ~0.025 μm2 in the rat. These results of the group reveal a robust difference in the biophysical properties of a well-defined synaptic connection of the cortical microcircuit of human and rodents.
Earlier experiments of the group performed in human brain slices suggested that individual human neurons can trigger temporally limited (15-20 ms) network events of alternating EPSPs and IPSPs following each other at ~200 Hz. Potentially similar, ultra-high frequency network events in the hippocampus are instrumental in a dialogue with the neocortex during memory formation, but the existence of transient ~200 Hz network events in the neocortex was not clear. The group developed a novel electrode holder and experimental procedure capable of stable (up to 40 mins) juxtacellular recordings from identified pyramidal cells and interneurons in completely anaesthesia free animals. Their recordings from neocortical layer II/III of freely behaving rats revealed field potential events at ripple and high gamma frequencies repeatedly occurring at troughs of spindle oscillations during sleep. Juxtacellular recordings identified subpopulations of fast spiking, parvalbumin containing basket cells with epochs of firing at ripple (~200 Hz) and high gamma (~120 Hz) frequencies detected during spindles and centered with millisecond precision at the trough of spindle waves in phase with field potential events but phase shifted relative to pyramidal cell firing. The results suggest that basket cell subpopulations are involved in spindle nested, high frequency network events which hypothetically provide repeatedly occurring neocortical temporal reference states potentially involved in mnemonic processes.