Projections of hippocampal CA2 pyramidal neurons: Distinct innervation patterns of CA2 compared to CA3 in rodents.

The hippocampus is a center for spatial and episodic memory formation in rodents. Understanding the composition of subregions and circuitry maps of the hippocampus is essential for elucidating the mechanism of memory formation and recall. For decades, the trisynaptic circuit (entorhinal cortex layer II-dentate gyrus - CA3-CA1) has been considered the neural network substrate responsible for learning and memory.

Single-Cell Labeling Strategies to Dissect Neuronal Structures and Local Functions.

The brain network consists of ten billion neurons and is the most complex structure in the universe. Understanding the structure of complex brain networks and neuronal functions is one of the main goals of modern neuroscience. Since the seminal invention of Golgi staining, single-cell labeling methods have been among the most potent approaches for dissecting neuronal structures and neural circuits.

Protocol for BATTLE-1EX: A High-Resolution Imaging Method to Visualize Whole Synaptic Structures and their Components in the Nervous System.

This protocol describes BATTLE-1EX, which is a combined method of BATTLE-1 and expansion microscopy to obtain high-resolution imaging of whole synaptic structures and their components of hippocampal neural circuits. BATTLE-1 uses two genetically engineered recombinase proteins and competition between two recombinases that can be independently titrated, resulting in a tunable proportion of mCherry+/YFP- and YFP+/mCherry- cells. As a combinational method, BATTLE-1EX has the potential to visualize and dissect whole synaptic structures in numerous regions in the brain.

BATTLE: Genetically Engineered Strategies for Split-Tunable Allocation of Multiple Transgenes in the Nervous System.

Elucidating fine architectures and functions of cellular and synaptic connections requires development of new flexible methods. Here, we created a concept called the "battle of transgenes," based on which we generated strategies using genetically engineered battles of multiple recombinases. The strategies enabled split-tunable allocation of multiple transgenes.

Island cells control temporal association memory.

Episodic memory requires associations of temporally discontiguous events. In the entorhinal-hippocampal network, temporal associations are driven by a direct pathway from layer III of the medial entorhinal cortex (MECIII) to the hippocampal CA1 region. However, the identification of neural circuits that regulate this association has remained unknown.

Cell type-specific genetic and optogenetic tools reveal hippocampal CA2 circuits.

The formation and recall of episodic memory requires precise information processing by the entorhinal-hippocampal network. For several decades, the trisynaptic circuit entorhinal cortex layer II (ECII)→dentate gyrus→CA3→CA1 and the monosynaptic circuit ECIII→CA1 have been considered the primary substrates of the network responsible for learning and memory. Circuits linked to another hippocampal region, CA2, have only recently come to light.

A local reduction in cortical GABAergic synapses after a loss of endogenous brain-derived neurotrophic factor, as revealed by single-cell gene knock-out method.

To address questions of whether brain-derived neurotrophic factor (BDNF) released from active excitatory neurons acts locally only on GABAergic presynaptic terminals contacting these neurons or generally also on GABAergic terminals contacting other inactive neurons, we developed a single-cell gene knock-out method in organotypic slice culture of visual cortex of floxed BDNF transgenic mice. A biolistic transfection of Cre recombinase with green fluorescence protein (GFP) plasmids to layer II/III of the cortex resulted in loss of BDNF in a single neuron or a small number of neurons, which expressed GFP at 13-14 d in vitro. Analysis with in situ hybridization and immunohistochemistry confirmed that neurons expressing GFP lacked BDNF mRNA and protein, respectively.

Difference in trafficking of brain-derived neurotrophic factor between axons and dendrites of cortical neurons, revealed by live-cell imaging.

Background: Brain-derived neurotrophic factor (BDNF), which is sorted into a regulated secretory pathway of neurons, is supposed to act retrogradely through dendrites on presynaptic neurons or anterogradely through axons on postsynaptic neurons. Depending on which is the case, the pattern and direction of trafficking of BDNF in dendrites and axons are expected to be different. To address this issue, we analyzed movements of green fluorescent protein (GFP)-tagged BDNF in axons and dendrites of living cortical neurons by time-lapse imaging.

Inhibitory but not excitatory cortical neurons require presynaptic brain-derived neurotrophic factor for dendritic development, as revealed by chimera cell culture.

To address questions of whether endogenous BDNF acts differentially on inhibitory and excitatory neurons, and through what routes, we used chimera culture of cerebral cortical neurons derived from BDNF-/- mice and another type of transgenic mice that express green fluorescence protein and BDNF. Presynaptic BDNF transferred to both types of neurons, GABA-synthesizing enzyme-positive and -negative neurons. The latter neurons were confirmed to be glutamatergic with immunocytochemistry.