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Perineuronal nets (PNNs) represent a highly condensed specialized form of brain extracellular matrix (ECM) enwrapping mostly parvalbumin-positive interneurons in the brain in a mesh-like fashion. PNNs not only regulate the onset and completion of the critical period during postnatal brain development, control cell excitability, and synaptic transmission but are also implicated in several brain disorders including schizophrenia. Holes in the perineuronal nets, harboring the synaptic contacts, along with hole-surrounding ECM barrier can be viewed as PNN compartmentalization units that might determine the properties of synapses and heterosynaptic communication. In this study, we developed a novel open-source script for Fiji (ImageJ) to semi-automatically quantify structural alterations of PNNs such as the number of PNN units, area, mean intensity of PNN marker expression in 2D and 3D, shape parameters of PNN units in the ketamine-treated Sprague-Dawley rat model of schizophrenia using high-resolution confocal microscopic images. We discovered that the mean intensity of ECM within PNN units is inversely correlated with the area and the perimeter of the PNN holes. The intensity, size, and shape of PNN units proved to be three major principal factors to describe their variability. Ketamine-treated rats had more numerous but smaller and less circular PNN units than control rats. These parameters allowed to correctly classify individual PNNs as derived from control or ketamine-treated groups with approximate to 85% reliability. Thus, the proposed multidimensional analysis of PNN units provided a robust and comprehensive morphometric fingerprinting of fine ECM structure abnormalities in the experimental model of schizophrenia.

The cell adhesion molecule, CHL1, like its close homologue L1, is important for normal brain development and function. In this study, we analysed the functional role of CHL1 in synaptic transmission in the CA1 region of the hippocampus using juvenile CHL1-deficient (CHL1–/–) and wild-type (CHL1+/+) mice. Inhibitory postsynaptic currents evoked in pyramidal cells by minimal stimulation of perisomatically projecting interneurons were increased in CHL1–/– mice compared with wild-type littermates. Also, long-term potentiation (LTP) at CA3–CA1 excitatory synapses was reduced under physiological conditions in CHL1-/– mice. This abnormality was abolished by application of a GABAA receptor antagonist, suggesting that enhanced inhibition is the cause of LTP impairment. Quantitative ultrastructural and immunohistochemical analyses revealed aberrations possibly related to the abnormally high inhibition observed in CHL1–/– mice. The length and linear density of active zones in symmetric synapses on pyramidal cell bodies, as well as number of perisomatic puncta containing inhibitory axonal markers were increased. Density and total number of parvalbumin-positive interneurons was also abnormally high. These observations and the finding that CA1 interneurons express CHL1 protein indicate that CHL1 is important for regulation of inhibitory synaptic transmission and interneuron populations in the postnatal brain. The observed enhancement of inhibitory transmission in CHL1–/– mice is in contrast to the previous finding of reduced inhibition in L1 deficient mice and indicates different functions of these two closely related molecules.

According to the local memory storage hypothesis, information about the tone-shock association in an auditory fear-conditioning paradigm is stored in synapses within the lateral amygdala. Thus, fear-conditioning-induced potentiation of auditory-evoked potentials in response to a conditioned stimulus (CS+, a series of short lasting tones; patterned tone) has been interpreted as an in vivo correlate of amygdaloid synaptic plasticity. Here, we re-examine the specificity of potentiation of auditory-evoked potentials in terms of (i) local confinement to the lateral amygdala, (ii) parameters of CS+ and (iii) influence of context, using a discriminatory fear-conditioning paradigm. Adult male C57BL/6J mice were implanted with recording electrodes aimed at the lateral amygdala, the CA1 region of the hippocampus and the neck muscles for simultaneous recordings of auditory-evoked potentials and startle responses. In a neutral context, auditory-evoked potentials within lateral amygdala and CA1 as well as startle and freezing responses to the CS+ were significantly potentiated following conditioning, as compared with pre-conditioning values and responses to a neutral stimulus (CSn; tone of different frequency). Potentiation was only evident if CS+ was presented as a uniform series but not if presented mixed with CSn. Accordingly, mice failed to show intensified freezing to a patterned tone if a single lasting tone of the same frequency served as CS+. Both CA1 and lateral amygdala auditory-evoked potentials were potentiated in response to CSn if presented in the conditioning context. These findings demonstrate that (i) potentiation of auditory-evoked potentials is not restricted to the lateral amygdala, (ii) both tone frequency and pattern of tone presentation are essential for proper CS+ recognition and (iii) contextual memory leads to a general potentiation of auditory-evoked potentials.

Perisomatic inhibition of pyramidal cells regulates efferent signalling from the hippocampus. The striking presence of HNK-1, a carbohydrate expressed by neural adhesion molecules, on perisomatic interneurons and around somata of CA1 pyramidal neurons led us to apply monoclonal HNK-1 antibodies to acute murine hippocampal slices. Injection of these antibodies decreased GABAA receptor-mediated perisomatic inhibitory postsynaptic currents (pIPSCs) but did not affect dendritic IPSCs or excitatory postsynaptic currents. The decrease in the mean amplitude of evoked pIPSCs by HNK-1 antibodies was accompanied by an increase in the coefficient of variation of pIPSC amplitude, number of failures and changes in frequency but not amplitude of miniature IPSCs, suggesting that HNK-1 antibodies reduced efficacy of evoked GABA release. HNK-1 antibodies did not affect pIPSCs in knock-out mice deficient in the extracellular matrix molecule tenascin-R which carries the HNK-1 carbohydrate as analysed by immunoblotting in synaptosomal fractions prepared from the CA1 region of the hippocampus. For control, HNK-1 antibody was applied to acute sections of mice deficient in the neural cell adhesion molecule NCAM, another potential carrier of HNK-1, and resulted in decrease of pIPSCs as observed in wild-type mice. Reduction in perisomatic inhibition is expected to promote induction of long-term potentiation (LTP) by increasing the level of depolarization during theta-burst stimulation. Indeed, LTP was increased by HNK-1 antibody applied before stimulation. Moreover, LTP was reduced by an HNK-1 peptide mimic, but not control peptide. These results provide first evidence that tenascin-R and its associated HNK-1 carbohydrate modulate perisomatic inhibition and synaptic plasticity in the hippocampus.

The distinction between pre- or postsynaptic expression of synaptic plasticity is difficult to make, unless the postsynaptic receptors can be investigated in isolation. We have studied single synaptic contacts in dissociated cultures of rat hippocampus. The reaction of postsynaptic receptor assemblies to the induction of synaptic plasticity was measured and compared with changes in the rate of spontaneous miniature excitatory postsynaptic currents (mEPSCs), which can reflect changes in the transmitter release mechanism. The response of a receptor assembly to locally applied exogenous glutamate was measured before and after synchronized application of glutamate and a train of postsynaptic depolarizations ('pairing'). Pairing induced a variety of changes: (i) the majority of the receptor assemblies showed no change in their response to glutamate before and after pairing; (ii) the postsynaptic current due to exogenous glutamate showed a rapid increase in five out of 26 cases. This was not due to changes in the single channel conductance; (iii) the rate of mEPSCs increased, if it had previously been below 25 Hz; (iv) the rate of mEPSCs decreased, if it had previously been above 25 Hz. Effects 2 and 3 were blocked by antagonists of NMDA receptors. These findings provide direct evidence for an increase of the number of glutamate receptors at a subset of the investigated postsynaptic sites during synaptic potentiation.