Optical investigation and manipulation constitute the core of biological experiments. Here, we introduce a new borosilicate glass material that contains the rare-earth ion terbium(III) (Tb3+), which emits green fluorescence upon blue light excitation, similar to green fluorescent protein (GFP), and thus is widely compatible with conventional biological research environments. Micropipettes made of Tb3+-doped glass allowed us to target GFP-labeled cells for single-cell electroporation, single-cell transcriptome analysis (Patch-seq), and patch-clamp recording under real-time fluorescence microscopic control. The glass also exhibited potent third harmonic generation upon infrared laser excitation and was usable for online optical targeting of fluorescently labeled neurons in the in vivo neocortex. Thus, Tb3+-doped glass simplifies many procedures in biological experiments.

The presubiculum, a subarea of the parahippocampal region, plays a critical role in spatial navigation and spatial representation. An outstanding aspect of presubicular spatial codes is head-direction selectivity of the firing of excitatory neurons, called head-direction cells. Head-direction selectivity emerges before eye-opening in rodents and is maintained in adulthood through neurophysiological interactions between excitatory and inhibitory neurons. Although the presubiculum has been physiologically profiled in terms of spatial representation during development, the histological characteristics of the developing presubiculum are poorly understood. We found that the expression of vesicular glutamate transporter 2 (VGluT2) could be used to delimit the superficial layers of the presubiculum, which was identified using an anterograde tracer injected into the anterior thalamic nucleus (ATN). Thus, we immunostained slices from mice ranging in age from neonates to adults using an antibody against VGluT2 to evaluate the VGluT2-positive area, which was identified as the superficial layers of the presubiculum, during development. We also immunostained the slices using antibodies against parvalbumin (PV) and somatostatin (SOM) and found that in the presubicular superficial layers, PV-positive neurons progressively increased in number during development, whereas SOM-positive neurons exhibited no increasing trend. In addition, we observed repeating patch structures in presubicular layer III from postnatal days 12. The abundant expression of VGluT2 suggests that the presubicular superficial layers are regulated primarily by VGluT2-mediated excitatory neurotransmission. Moreover, developmental changes in the densities of PV- and SOM-positive interneurons and the emergence of the VGluT2-positive patch structures during adolescence may be associated with the functional development of spatial codes in the superficial layers of the presubiculum.

The effect of excitatory synaptic input on the excitation of the cell body is believed to vary depending on where and when the synaptic activation occurs in dendritic trees and the spatiotemporal modulation by inhibitory synaptic input. However, few studies have examined how individual synaptic inputs influence the excitability of the cell body in spontaneously active neuronal networks mainly because of the lack of an appropriate method. We developed a calcium imaging technique that monitors synaptic inputs to hundreds of spines from a single neuron with millisecond resolution in combination with whole-cell patch-clamp recordings of somatic excitation. In rat hippocampal CA3 pyramidal neurons ex vivo, a fraction of the excitatory synaptic inputs were not detectable in the cell body against background noise. These synaptic inputs partially restored their somatic impact when a GABAA receptor blocker was intracellularly perfused. Thus, GABAergic inhibition reduces the influence of some excitatory synaptic inputs on the somatic excitability. Numerical simulation using a single neuron model demonstrates that the timing and locus of a dendritic GABAergic input are critical to exert this effect. Moreover, logistic regression analyses suggest that the GABAergic inputs sectionalize spine activity; that is, only some subsets of synchronous synaptic activity seemed to be preferably passed to the cell body. Thus, dendrites actively sift inputs from specific presynaptic cell assemblies.

In some models, animals approach aversive stimuli more than those housed in an enriched environment. Here, we found that male mice in an impoverished and unstimulating (i.e., boring) chamber without toys sought aversive air puffs more often than those in an enriched chamber. Using this animal model, we identified the insular cortex as a regulator of aversion-seeking behavior. Activation and inhibition of the insular cortex increased and decreased the frequencies of air-puff self-stimulation, respectively, and the firing patterns of insular neuron ensembles predicted the self-stimulation timing. Dopamine levels in the ventrolateral striatum decreased with passive air puffs but increased with actively sought puffs. Around 20% of mice developed intense self-stimulation despite being offered toys, which was prevented by administering opioid receptor antagonists. This study establishes a basis for comprehending the neural underpinnings of usually avoided stimulus-seeking behaviors.