Oyabu, K. | bioRxiv Science

bioRxiv2023

Location analysis of presynaptically active and silent synapses in single-cultured hippocampal neurons

A morphologically present but non-functioning synapse is termed a silent synapse. Silent synapses are categorized into "postsynaptically silent synapses," where AMPA receptors are either absent or non-functional, and "presynaptically silent synapses," where neurotransmitters cannot be released from nerve terminals. The presence of presynaptically silent synapses remains enigmatic, and their physiological significance is highly intriguing. In this study, we examined the distribution and developmental changes of presynaptically active and silent synapses in individual neurons. Our findings show a gradual increase in the number of excitatory synapses, along with a corresponding decrease in the percentage of presynaptically silent synapses during neuronal development. To pinpoint the distribution of presynaptically active and silent synapses, i.e., their positional information, we enhanced the traditional Sholl analysis and introduced a novel method termed "donut analysis." Our results indicate that the distribution of presynaptically silent synapses within a single neuron does not exhibit a distinct pattern during synapse development in different areas. However, irrespective of neuronal development, the proportion of presynaptically silent synapses tends to rise as the projection site moves farther from the cell body, suggesting that synapses near the cell body may exhibit higher synaptic transmission efficiency. This study represents the first observation of changes in the distribution of presynaptically active and silent synapses within a single neuron. Additionally, we propose that donut analysis can serve as a valuable analytical tool for evaluating synaptic positional information. Scope statementA morphologically present but non-functioning synapse is termed a silent synapse. The presence of presynaptically silent synapses remains enigmatic, and their physiological significance is highly intriguing. This study focused on the distribution and developmental changes of presynaptically active and silent synapses in individual neurons. To pinpoint the distribution of presynaptically active and silent synapses, we enhanced the traditional Sholl analysis and introduced a novel method termed "donut analysis." We found that the distribution of presynaptically silent synapses within a single neuron does not exhibit a distinct pattern during synapse development in different areas. However, irrespective of neuronal development, the proportion of presynaptically silent synapses tends to rise as the projection site moves farther from the cell body. This is a new paper that applies "Sholl analysis," a method invented 70 years ago that is now the gold standard of morphological analysis of the single neuron. This study represents the first observation of changes in the distribution of presynaptically active and silent synapses within a single neuron. Additionally, we propose that donut analysis can serve as a valuable analytical tool for evaluating synaptic positional information for the design of "synaptic maps" in neural circuits.

neuroscience↗

bioRxiv2022

Isolation of bat sarbecoviruses of SARS-CoV-2 clade, Japan

Betacoronaviruses have caused 3 outbreaks in the past 2 decades. SARS-CoV-2, in particular, has caused a serious pandemic. As the betacoronaviruses are considered to originate from bats, surveillance of bat betacoronaviruses is crucial for understanding the mechanism of cross-species transition and potential for future outbreaks. We previously detected and characterized a SARS-CoV-2-related sarbecovirus, Rc-o319, from Rhinolophus cornutus in Japan. Here, we detected several bat sarbecoviruses of the SARS-CoV-2 clade from R. cornutus in multiple locations in Japan, and successfully isolated them using Vero/TMPRSS2 cells stably expressing R. cornutus ACE2 (Vero-RcACE2). The coding sequences of S1 region varied among isolates, whereas other genetic regions were highly conserved. Isolates were efficiently grown in Vero-RcACE2 cells, but did not replicate in Vero/TMPRSS2 cells stably expressing human ACE2, suggesting a narrow host range. Further long-term epidemiological studies of sarbecoviruses in wildlife are expected to facilitate the assessment of the risk of their spillover potential.

microbiology↗

bioRxiv2020

Presynaptically silent synapses are modulated by the density of surrounding astrocytes

The astrocyte, a major glial cell type, is involved in formation and maturation of synapses, and thus contributes to sustainable synaptic transmission between neurons. Given that the animals in the higher phylogenetic tree have brains with higher density of glial cells with respect to neurons, there is a possibility that the relative astrocytic density directly influences synaptic transmission. However, the notion has not been tested thoroughly. Here we addressed it, by using a primary culture preparation where single hippocampal neurons are surrounded by a variable but countable number of cortical astrocytes in dot-patterned microislands, and recording synaptic transmission by patch-clamp electrophysiology. Neurons with a higher astrocytic density showed a higher amplitude of evoked excitatory postsynaptic current (EPSC) than that of neurons with a lower astrocytic density. The size of readily releasable pool of synaptic vesicles per neuron was significantly higher. The frequency of spontaneous synaptic transmission (miniature EPSC) was higher, but the amplitude was unchanged. The number of morphologically identified glutamatergic synapses was unchanged, but the number of functional ones was increased, indicating a lower ratio of presynaptically silent synapses. Taken together, the higher astrocytic density enhanced excitatory synaptic transmission by increasing the number of functional synapses through presynaptic un-silencing.

neuroscience↗