Structure of Excitatory Synapses and GABAA Receptor Localization at Inhibitory Synapses Are Regulated by Neuroplastin-65

Abstract

Formation, maintenance, and activity of excitatory and inhibitory synapses are essential for neuronal network function. Cell adhesion molecules (CAMs) are crucially involved in these processes. The CAM neuroplastin-65 (Np65) highly expressed during periods of synapse formation and stabilization is present at the pre- and postsynaptic membranes. Np65 can translocate into synapses in response to electrical stimulation and it interacts with subtypes of GABAA receptors in inhibitory synapses. Here, we report that in the murine hippocampus and in hippocampal primary culture, neurons of the CA1 region and the dentate gyrus (DG) express high Np65 levels, whereas expression in CA3 neurons is lower. In neuroplastin-deficient (Np(-/-)) mice the number of excitatory synapses in CA1 and DG, but not CA3 regions is reduced. Notably this picture is mirrored in mature Np(-/-) hippocampal cultures or in mature CA1 and DG wild-type (Np(+/+)) neurons treated with a function-blocking recombinant Np65-Fc extracellular fragment. Although the number of GABAergic synapses was unchanged in Np(-/-) neurons or in mature Np65-Fc-treated Np(+/+) neurons, the ratio of excitatory to inhibitory synapses was significantly lower in Np(-/-) cultures. Furthermore, GABAA receptor composition was altered at inhibitory synapses in Np(-/-) neurons as the α1 to α2 GABAA receptor subunit ratio was increased. Changes of excitatory and inhibitory synaptic function in Np(-/-) neurons were confirmed evaluating the presynaptic release function and using patch clamp recording. These data demonstrate that Np65 is an important regulator of the number and function of synapses in the hippocampus.

The Neuroplastin adhesion molecules: key regulators of neuronal plasticity and synaptic function

“…Recent studies using hippocampal neurones derived from mice in which expression of the NPTN gene is disrupted show that both the structure and function of glutamatergic and the function of GABAergic synapses in hippocampal CA1 and DG regions are regulated by Np65 (Herrera‐Molina et al . ). Western blotting and immunocytochemistry confirm that there is no detectable Neuroplastin in brain in Neuroplastin‐deficient mice.…”

“…Electrophysiological studies showed that both the amplitude and frequency of miniature excitatory postsynaptic currents, mEPSCs, are reduced by some 33% as a result of Neuroplastin deficiency (Herrera‐Molina et al . ). In contrast, GABAergic synapses showed no effect of Np65 deficiency on either number or matching of pre‐and postsynaptic elements.…”

“…Rat hippocampal neurones were cultured for 28 DIV (Kaech and Banker ) and live stained for Np65 (red) using an anti‐Np65 isoform‐specific antibody according to published protocols (Herrera‐Molina et al . ). Cells were then fixed, permeabilized and stained with antibodies against pre‐(Synapsin I, blue) and postsynaptic (Homer, green) markers followed by secondary antibodies.…”

“…A recent study comparing the binding of Np65Fc to hippocampal neurones from wild‐type and Neuroplastin‐deficient mice confirms that the recombinant protein does indeed bind to endogenous Np65 molecules (Herrera‐Molina et al . ).…”

The balancing act of GABAergic synapse organizers

“…Knockdown of neuroplastin-65 decreases GABA A receptors at synaptic sites, suggesting that it is essential for the maintenance of a subset of GABA A receptors [45]. In neuroplastin-65-deficient mice, both excitatory and inhibitory synapses are influenced in CA1 and the DG of the hippocampus [46], and GABA A receptor composition is drastically altered in neuroplastin-65-KO mice [46].…”