Diffuse perineuronal nets and modified pyramidal cells immunoreactive for glutamate and the GABAA receptor α1 subunit form a unique entity in rat cerebral cortex
Introduction
Perineuronal nets (PNs) are specialized accumulations of extracellular matrix components around certain types of neurons in cortical and subcortical brain regions (for reviews, see Celio and Blümcke, 1994, Carlson and Hockfield, 1996, Celio et al., 1998). More than one century after their first description by Golgi (1893) as “pericellular nets”, the functional significance of this exceptional neuronal microenvironment is still a matter of debate. Several aspects have been discussed, such as the involvement in synaptic stabilization and in limitation of synaptic plasticity Fox and Caterson, 2002, Hockfield et al., 1990, Lander et al., 1997, Pizzorusso et al., 2002, a neuroprotective role Brückner et al., 1999, Okamoto et al., 1994, Schüppel et al., 2002, and the support of ion homeostasis around highly active types of neurons Brückner et al., 1993, Brückner et al., 1996a, Brückner et al., 1996b, Härtig et al., 1999, Härtig et al., 2001, Hobohm et al., 1998.
PNs consist of polyanionic chondroitin sulfate proteoglycans (CSPG) and associated molecules such as hyaluronic acid and tenascins Brückner et al., 1993, Brückner et al., 2000, Haghihara et al., 1999, Köppe et al., 1997, Matsui et al., 1999, Matthews et al., 2002. They ensheath the perikaryon, the proximal parts of dendrites, the axon initial segment, and all presynaptic boutons attached to these structures Brückner et al., 1993, Brückner et al., 1996b, Hendry et al., 1988. Adjacent astrocytic processes usually contact the matrix material, forming a specialized neuron-glia interface Blümcke et al., 1995, Brauer et al., 1984, Brückner et al., 1993, Brückner et al., 1996b, Derouiche et al., 1996.
The neocortex contains high numbers of PNs in motor and primary sensory areas, whereas they are less numerous in the association and limbic cortices Brückner et al., 1994, Brückner et al., 1999, Hausen et al., 1996, McGuire et al., 1989. Cortical PNs are mainly associated with GABAergic interneurons Brückner et al., 1994, Mulligan et al., 1989, Naegele et al., 1988, Nagakawa et al., 1986, but they were also detected around certain pyramidal neurons Brückner et al., 1999, Härtig et al., 1999, Hausen et al., 1996, Ohyama and Ojima, 1997. Most of the net-associated interneurons express the calcium-binding protein parvalbumin Härtig et al., 1992, Härtig et al., 1994, Kosaka et al., 1990, Morino-Wannier et al., 1992 and the voltage-gated potassium channel subunit Kv3.1b (Härtig et al., 1999), which is supposed to be a prerequisite for the generation of high firing rates (see, e.g., Chow et al., 1999, Gan and Kaczmarek, 1998). Fast synaptic inhibition is mediated mainly by activation of GABAA receptor channels in the mammalian CNS Macdonald and Olsen, 1994, Whiting et al., 1995. In rat neocortex, the α1 subunit of GABAA receptors is most abundant in interneurons (Fritschy and Möhler, 1995). Cortical nonpyramidal interneurons with PNs are known to be characterized by a multipolar morphology and aspiny or sparsely spinous dendrites. However, only few of the net-associated neurons have been identified by intracellular injections, some of them being described as large basket cells Naegele and Katz, 1990, Ojima, 1993.
The structure of the PNs appears to be adapted to the type of neuron ensheathed suggesting neuron-specific functions of the perineuronal extracellular matrix Brückner et al., 1993, Brückner et al., 1996b. In cerebral cortex, the inhibitory interneurons are associated with clearly contoured, lattice-like PNs, whereas pyramidal PNs possess a very faint structure Härtig et al., 1999, Ohyama and Ojima, 1997, Ojima et al., 1998.
The present study was undertaken to characterize the population of neocortical neurons surrounded by diffuse PNs, a distinct third class of PNs, which has not been described so far. We combine the N-acetylgalactosamine-binding Wisteria floribunda agglutinin (WFA) as an established marker for PNs (Härtig et al., 1992) with immunocytochemical staining, retrograde tracing, and intracellular injection of net-associated neurons in rat parietal cortex.
Section snippets
Animals
All Wistar rats used in this study were treated in agreement with the German law on the use of laboratory animals, and followed the ethical guidelines of the Laboratory Animal Care and Use Committee at the University of Leipzig.
Double fluorescence staining
Eight adult Wistar rats (200–300 g) of either sex were anaesthetized with ketamine (200 mg/kg) and then transcardially perfused with saline followed by the fixative (4% paraformaldehyde and 0.1% glutaraldehyde in 0.1 M phosphate buffer, pH 7.4). Glutaraldehyde was
Types of perineuronal nets in rat parietal cortex
Three types of WFA-stained PNs could be distinguished by their different morphological structure: nonpyramidal, pyramidal, and diffuse PNs (Fig. 1). Nonpyramidal PNs, described in many previous studies, were the largest group of PNs in rat parietal cortex (67% in area Par1). These nets surrounded the soma, proximal parts of dendrites and the axon initial segment of nonpyramidal neurons with a clearly contoured matrix sheath of approximately 1- to 2-μm thickness (Fig. 1A). Almost one-third of
Three types of perineuronal nets in rat cerebral cortex
The present study confirmed and extended results of previous investigations indicating that the extracellular matrix is specifically adapted to the architecture of brain regions and to distinct types of neurons (for review, see Carlson and Hockfield, 1996). In the mammalian cerebral cortex, subclasses of interneurons and pyramidal cells have been shown to be associated with PNs in area-specific proportions Brückner et al., 1994, Brückner et al., 1999, Hendry et al., 1988, McGuire et al., 1989,
Acknowledgements
The work was supported by the Deutsche Forschungsgemeinschaft (Graduiertenkolleg Intercell, Leipzig). We wish to thank Mrs. M. Schmidt (Leipzig) for her excellent technical assistance and J.-M. Fritschy (Zürich), W.H. Oertel (München), and O.P. Ottersen (Oslo) for their antisera.
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2021, Journal of Chemical NeuroanatomyCitation Excerpt :A portion of PNs is not detectable by WFA, but immunopositive for their main CSPG component aggrecan, as reported for instance for the murine hippocampus (Yamada and Jinno, 2017), whereas WFA-stainable, aggrecan-immunonegative PNs were found in the mouse entorhinal cortex (Ueno et al., 2018) and are also demonstrated in the present study (see Fig. 4d). While in the neocortex and numerous further brain regions WFA-binding sites surround predominantly fast-firing, parvalbumin-containing GABAergic neurons (Härtig et al., 1999; Wingert and Sorg, 2021), a considerable portion of nets enwraps neurons containing other neurotransmitters (Wegner et al., 2003). Notably, net-bearing cholinergic neurons were detected in the human nucleus abducens (Horn et al., 2018) and in the rat spinal cord (Irvine and Kwok, 2018).