Maturation Of Perineuronal Nets Research Articles

Net gain and loss: influence of natural rewards and drugs of abuse on perineuronal nets.

Overindulgence, excessive consumption, and a pattern of compulsive use of natural rewards, such as certain foods or drugs of abuse, may result in the development of obesity or substance use disorder, respectively. Natural rewards and drugs of abuse can trigger similar changes in the neurobiological substrates that drive food- and drug-seeking behaviors. This review examines the impact natural rewards and drugs of abuse have on perineuronal nets (PNNs). PNNs are specialized extracellular matrix structures that ensheathe certain neurons during development over the critical period to provide synaptic stabilization and a protective microenvironment for the cells they surround. This review also analyzes how natural rewards and drugs of abuse impact the density and maturation of PNNs within reward-associated circuitry of the brain, which may contribute to maladaptive food- and drug-seeking behaviors. Finally, we evaluate the relatively few studies that have degraded PNNs to perturb reward-seeking behaviors. Taken together, this review sheds light on the complex way PNNs are regulated by natural rewards and drugs and highlights a need for future studies to delineate the molecular mechanisms that underlie the modification and maintenance of PNNs following exposure to rewarding stimuli.

Mirtazapine treatment in a young female mouse model of Rett syndrome identifies time windows for the rescue of early phenotypes

Rett Syndrome (RTT) is a rare X-linked neurodevelopmental disorder, mainly caused by mutations in the MECP2 gene. Reduction in monoamine levels in RTT patients and mouse models suggested the possibility to rescue clinical phenotypes through antidepressants. Accordingly, we tested mirtazapine (MTZ), a noradrenergic and specific-serotonergic tetracyclic antidepressant (NaSSA). In previous studies, we showed high tolerability and significant positive effects of MTZ in male Mecp21m1.1Bird-knock-out mice, adult female Mecp2tm1.1Bird-heterozygous (Mecp2+/−) mice, and adult female RTT patients. However, it remained to explore MTZ efficacy in female Mecp2+/− mice at young ages. As RTT-like phenotypes in young Mecp2+/− mice have been less investigated, we carried out a behavioural characterization to analyze Mecp2+/− mice in “early adolescence” (6 weeks) and “young adulthood” (11 weeks) and identified several progressive phenotypes. Then, we evaluated the effects of either a 15- or a 30-day MTZ treatment on body weight and impaired motor behaviours in 11-week-old Mecp2+/− mice. Finally, since defective cortical development is a hallmark of RTT, we performed a histological study on the maturation of perineuronal nets (PNNs) and parvalbuminergic (PV) neurons in the primary motor cortex. The 30-day MTZ treatment was more effective than the shorter 15-day treatment, leading to the significant rescue of body weight, hindlimb clasping and motor learning in the accelerating rotarod test. Behavioural improvement was associated with normalized PV immunoreactivity levels and PNN thickness. These results support the use of MTZ as a new potential treatment for adolescent girls affected by RTT and suggest a possible mechanism of action.

Enzymatic Degradation of Cortical Perineuronal Nets Reverses GABAergic Interneuron Maturation

Perineuronal nets (PNNs) are specialised extracellular matrix structures which preferentially enwrap fast-spiking (FS) parvalbumin interneurons and have diverse roles in the cortex. PNN maturation coincides with closure of the critical period of cortical plasticity. We have previously demonstrated that BDNF accelerates interneuron development in a c-Jun-NH2-terminal kinase (JNK)–dependent manner, which may involve upstream thousand-and-one amino acid kinase 2 (TAOK2). Chondroitinase-ABC (ChABC) enzymatic digestion of PNNs reportedly reactivates ‘juvenile-like’ plasticity in the adult CNS. However, the mechanisms involved are unclear. We show that ChABC produces an immature molecular phenotype in cultured cortical neurons, corresponding to the phenotype prior to critical period closure. ChABC produced different patterns of PNN-related, GABAergic and immediate early (IE) gene expression than well-characterised modulators of mature plasticity and network activity (GABAA-R antagonist, bicuculline, and sodium-channel blocker, tetrodotoxin (TTX)). ChABC downregulated JNK activity, while this was upregulated by bicuculline. Bicuculline, but not ChABC, upregulated Bdnf expression and ERK activity. Furthermore, we found that BDNF upregulation of semaphorin-3A and IE genes was TAOK mediated. Our data suggest that ChABC heightens structural flexibility and network disinhibition, potentially contributing to ‘juvenile-like’ plasticity. The molecular phenotype appears to be distinct from heightened mature synaptic plasticity and could relate to JNK signalling. Finally, we highlight that BDNF regulation of plasticity and PNNs involves TAOK signalling.

Child abuse associates with increased recruitment of perineuronal nets in the ventromedial prefrontal cortex: a possible implication of oligodendrocyte progenitor cells

Child abuse (CA) is a strong predictor of psychopathologies and suicide, altering normal trajectories of brain development in areas closely linked to emotional responses such as the prefrontal cortex (PFC). Yet, the cellular underpinnings of these enduring effects are unclear. Childhood and adolescence are marked by the protracted formation of perineuronal nets (PNNs), which orchestrate the closure of developmental windows of cortical plasticity by regulating the functional integration of parvalbumin interneurons into neuronal circuits. Using well-characterized post-mortem brain samples, we show that a history of CA is specifically associated with increased densities and morphological complexity of WFL-labeled PNNs in the ventromedial PFC (BA11/12), possibly suggesting increased recruitment and maturation of PNNs. Through single-nucleus sequencing and fluorescent in situ hybridization, we found that the expression of canonical components of PNNs is enriched in oligodendrocyte progenitor cells (OPCs), and that they are upregulated in CA victims. These correlational findings suggest that early-life adversity may lead to persistent patterns of maladaptive behaviors by reducing the neuroplasticity of cortical circuits through the enhancement of developmental OPC-mediated PNN formation.

Perineuronal Nets and Metal Cation Concentrations in the Microenvironments of Fast-Spiking, Parvalbumin-Expressing GABAergic Interneurons: Relevance to Neurodevelopment and Neurodevelopmental Disorders.

Because of their abilities to catalyze generation of toxic free radical species, free concentrations of the redox reactive metals iron and copper are highly regulated. Importantly, desired neurobiological effects of these redox reactive metal cations occur within very narrow ranges of their local concentrations. For example, synaptic release of free copper acts locally to modulate NMDA receptor-mediated neurotransmission. Moreover, within the developing brain, iron is critical to hippocampal maturation and the differentiation of parvalbumin-expressing neurons, whose soma and dendrites are surrounded by perineuronal nets (PNNs). The PNNs are a specialized component of brain extracellular matrix, whose polyanionic character supports the fast-spiking electrophysiological properties of these parvalbumin-expressing GABAergic interneurons. In addition to binding cations and creation of the Donnan equilibrium that support the fast-spiking properties of this subset of interneurons, the complex architecture of PNNs also binds metal cations, which may serve a protective function against oxidative damage, especially of these fast-spiking neurons. Data suggest that pathological disturbance of the population of fast-spiking, parvalbumin-expressing GABAergic inhibitory interneurons occur in at least some clinical presentations, which leads to disruption of the synchronous oscillatory output of assemblies of pyramidal neurons. Increased expression of the GluN2A NMDA receptor subunit on parvalbumin-expressing interneurons is linked to functional maturation of both these neurons and the perineuronal nets that surround them. Disruption of GluN2A expression shows increased susceptibility to oxidative stress, reflected in redox dysregulation and delayed maturation of PNNs. This may be especially relevant to neurodevelopmental disorders, including autism spectrum disorder. Conceivably, binding of metal redox reactive cations by the perineuronal net helps to maintain safe local concentrations, and also serves as a reservoir buffering against second-to-second fluctuations in their concentrations outside of a narrow physiological range.

Perineuronal net degradation rescues CA2 plasticity in a mouse model of Rett syndrome.

Perineuronal nets (PNNs), a specialized form of extracellular matrix, are abnormal in the brains of people with Rett syndrome (RTT). We previously reported that PNNs function to restrict synaptic plasticity in hippocampal area CA2, which is unusually resistant to long-term potentiation (LTP) and has been linked to social learning in mice. Here we report that PNNs appear elevated in area CA2 of the hippocampus of an individual with RTT and that PNNs develop precociously and remain elevated in area CA2 of a mouse model of RTT (Mecp2-null). Further, we provide evidence that LTP could be induced at CA2 synapses prior to PNN maturation (postnatal day 8-11) in wild-type mice and that this window of plasticity was prematurely restricted at CA2 synapses in Mecp2-null mice. Degrading PNNs in Mecp2-null hippocampus was sufficient to rescue the premature disruption of CA2 plasticity. We identified several molecular targets that were altered in the developing Mecp2-null hippocampus that may explain aberrant PNNs and CA2 plasticity, and we discovered that CA2 PNNs are negatively regulated by neuronal activity. Collectively, our findings demonstrate that CA2 PNN development is regulated by Mecp2 and identify a window of hippocampal plasticity that is disrupted in a mouse model of RTT.

Perineuronal net abnormalities in Slc13a4+/− mice are rescued by postnatal administration of N-acetylcysteine

Disruptions to either sulfate supply or sulfation enzymes can affect brain development and have long-lasting effects on brain function, yet our understanding of the molecular mechanisms governing this are incomplete. Perineuronal nets (PNNs) are highly sulfated, specialized extracellular matrix structures that regulate the maturation of synaptic connections and neuronal plasticity. We have previously shown that mice heterozygous for the brain sulfate transporter Slc13a4 have abnormal social interactions, memory, exploratory behaviors, stress and anxiety of postnatal origin, pointing to potential deficits in PNN biology, and implicate SLC13A4 as a critical factor required for regulating normal synaptic connectivity and function. Here, we sought to investigate aberrant PNN formation as a potential mechanism contributing to the functional deficits displayed by Slc13a4+/− mice. Following social interactions, we reveal reduced neuronal activation in the somatosensory cortex of Slc13a4+/− mice, and altered inhibitory and excitatory postsynaptic currents. In line with this, we found a reduction in parvalbumin-expressing neurons decorated with PNNs, as well as reduced expression of markers for PNN maturation. Finally, we reveal that postnatal administration of N-acetylcysteine prevented PNN abnormalities from manifesting in Slc13a4+/− adult animals. Collectively, these data highlight a central role for postnatal SLC13A4 in normal PNN formation, circuit function and subsequent animal behavior.

Chondroitin sulfate expression around spinal motoneurons during postnatal development in rats

Perineuronal nets are extracellular matrix structures that surround neuronal cell bodies and their proximal dendrites in the central nervous system. Chondroitin sulfate proteoglycans, which contain chondroitin sulfates (CSs) are major components of perineuronal nets. CSs are considered to have inhibitory roles in neural plasticity, although the effects differ according to their sulfation pattern. In the present study, we investigated the expression of the CS subtypes CS-A and CS-C surrounding spinal motoneurons in different postnatal periods to explore the potential influence of altered CS sulfation patterns on spinal development.CS-A-positive structures were observed around motoneurons in the cervical, thoracic, and lumbar segments as early as postnatal day (P) 5. Most motoneurons were covered with CS-A-positive structures during the first 2 postnatal weeks. The percentage of motoneurons covered with CS-A-positive structures decreased after P20, becoming lower than 70% in the cervical, and lumber segments after P35. CS-C-positive structures were occasionally observed around motoneurons during the first 2 postnatal weeks. The percentage of motoneurons covered with CS-C-positive structures increased after P20, becoming significantly higher after P25 than before P20. The expression pattern of Wisteria Floribunda agglutinin-positive structures around motoneurons was similar to that of the CS-C–positive structures.The present findings revealed that CS-A and CS-C are differentially expressed in the extracellular matrix surrounding motoneurons. The altered sulfation pattern with increased CS-C expression is associated with the maturation of perineuronal nets and might lead to changes in the motoneuron plasticity.

Does maternal separation accelerate maturation of perineuronal nets and parvalbumin-containing inhibitory interneurons in male and female rats?

Early life adversity impacts on a range of emotional, cognitive, and psychological processes. A recent theoretical model suggests that at least some of these effects are due to accelerated maturation of specific physiological systems and/or neural circuits. For example, maternal separation (MS), a model of early life adversity in rodents, accelerates maturation of memory systems, and here we examined its impact on maturation of perineuronal nets (PNNs) and parvalbumin (PV)-containing inhibitory interneurons. PNNs are specialized extracellular matrix structures suggested to be involved in stabilizing long-term memories and in the closure of a sensitive period in memory development. PV-containing inhibitory interneurons are the type of cell that PNNs preferentially surround, and are also thought to be involved in memory. In Experiment 1, with male rats, there was an increase in PNNs in both the amygdala and prefrontal cortex with age from infancy to juvenility. Contrary to prediction, MS had no impact on either PNN or PV expression. The same pattern was observed in female rats in Experiment 2. Taken together, these data show that the early maturation of memory in MS infants is not due to an accelerated maturation of PNNs or PV-containing cells in either the amygdala or prefrontal cortex.

BDNF and JNK Signaling Modulate Cortical Interneuron and Perineuronal Net Development: Implications for Schizophrenia-Linked 16p11.2 Duplication Syndrome.

Schizophrenia (SZ) is a neurodevelopmental disorder caused by the interaction of genetic and environmental risk factors. One of the strongest genetic risk variants is duplication (DUP) of chr.16p11.2. SZ is characterized by cortical gamma-amino-butyric acid (GABA)ergic interneuron dysfunction and disruption to surrounding extracellular matrix structures, perineuronal nets (PNNs). Developmental maturation of GABAergic interneurons, and also the resulting closure of the critical period of cortical plasticity, is regulated by brain-derived neurotrophic factor (BDNF), although the mechanisms involved are unknown. Here, we show that BDNF promotes GABAergic interneuron and PNN maturation through JNK signaling. In mice reproducing the 16p11.2 DUP, where the JNK upstream activator Taok2 is overexpressed, we find that JNK is overactive and there are developmental abnormalities in PNNs, which persist into adulthood. Prefrontal cortex parvalbumin (PVB) expression is reduced, while PNN intensity is increased. Additionally, we report a unique role for TAOK2 signaling in the regulation of PVB interneurons. Our work implicates TAOK2-JNK signaling in cortical interneuron and PNN development, and in the responses to BDNF. It also demonstrates that over-activation of this pathway in conditions associated with SZ risk causes long-lasting disruption in cortical interneurons.

Accelerated Hyper-Maturation of Parvalbumin Circuits in the Absence of MeCP2.

Methyl-CpG-binding protein 2 (MeCP2) mutations are the primary cause of Rett syndrome, a severe neurodevelopmental disorder. Cortical parvalbumin GABAergic interneurons (PV) make exuberant somatic connections onto pyramidal cells in the visual cortex of Mecp2-deficient mice, which contributes to silencing neuronal cortical circuits. This phenotype can be rescued independently of Mecp2 by environmental, pharmacological, and genetic manipulation. It remains unknown how Mecp2 mutation can result in abnormal inhibitory circuit refinement. In the present manuscript, we examined the development of GABAergic circuits in the primary visual cortex of Mecp2-deficient mice. We identified that PV circuits were the only GABAergic interneurons to be upregulated, while other interneurons were downregulated. Acceleration of PV cell maturation was accompanied by increased PV cells engulfment by perineuronal nets (PNNs) and by an increase of PV cellular and PNN structural complexity. Interestingly, selective deletion of Mecp2 from PV cells was sufficient to drive increased structure complexity of PNN. Moreover, the accelerated PV and PNN maturation was recapitulated in organotypic cultures. Our results identify a specific timeline of disruption of GABAergic circuits in the absence of Mecp2, indicating a possible cell-autonomous role of MeCP2 in the formation of PV cellular arbors and PNN structures in the visual cortex.

Perineuronal Nets Restrict the Induction of Long-Term Depression in the Mouse Hippocampal CA1 Region.

Long-term depression (LTD) of synaptic efficacy is widely regarded as a cellular basis of learning and memory. The magnitude of hippocampal CA1 LTD induced by low-frequency stimulation (LFS) declines with age, but the mechanisms involved remain poorly understood. Perineuronal nets (PNNs) are specialized extracellular matrix structures that function in dampening synaptic plasticity during postnatal development, suggesting that PNN formation may restrict LTD induction in the adult hippocampus. Here, we show that PNNs tightly enwrap a subpopulation of parvalbumin (PV) interneurons in the hippocampal CA1 region and enzymatic removal of PNNs with the chondroitinase ABC alters the excitatory/inhibitory synaptic balance toward more excitation and restores the ability of LFS to induce an N-methyl-D-aspartate receptor-dependent LTD at Schaffer collateral-CA1 synapses in slices from male adult mice. Early interference with depolarizing GABA with Na+-K+-2Cl- cotransporter inhibitor bumetanide impairs the maturation of PNNs and enhances LTD induction. These results provide novel insights into a previously unrecognized role for PNNs around PV interneurons in restricting long-term synaptic plasticity at excitatory synapses on hippocampal CA1 neurons in adulthood.

P.1.010 - Brain-derived neurotrophic factor enhances glutamic acid decarboxylase and perineuronal net maturation in mouse primary cortical cultures

Schizophrenia is a debilitating mental illness which affects approximately 1% of the global population, and is characterised by positive (e.g. hallucinations) and negative (e.g. anhedonia) symptoms, combined with cognitive deficits. Evidence suggests diminished cortical inhibitory drive as a major contributing factor in schizophrenia pathophysiology. Reductions in glutamic acid decarboxylase (GAD65/67) and deficits in the extracellular matrix structures, perineuronal nets (PNNs), which enwrap parvalbumin-containing interneurons have been observed in patients [1,2]. Members of the c-Jun N-Terminal kinase (JNK) family of mitogen-activated protein kinases (MAPKs) and their upstream kinase MKK7 have been implicated in neocortical development. Interestingly, MAP2K7 (encoding MKK7) has been implicated as a candidate risk gene for schizophrenia [3]. However, the impact of disrupted MAP2K7-JNK signalling on GABAergic interneuron development remains elusive. Reduced levels of brain-derived neurotrophic factor (BDNF) have also been found in patients [4]. Previous research highlights BDNF as a promoter of GABAergic maturation in hippocampal neurons [5]. Consequently, altered BDNF activity could result in abnormal cortical circuitry development and contribute to the cortical neuronal dysfunction underlying schizophrenia. Here we study the effects of BDNF on the development of primary cortical GABAergic interneurons and surrounding PNNs along with the possible involvement of JNK signalling. Mouse primary cortical neuronal cultures were treated at 7 days in vitro with either vehicle, BDNF (50 ng/ml), JNK inhibitor (SP600125, 5 μM) or BDNF+JNK inhibitor. Subsequently, immunofluorescent-cytochemistry (ICC) was performed. PNN development was assessed using Wisteria floribunda agglutinin (WFA) lectin which labels interneuron PNNs. Cultures were double-labelled with GAD65/67. Normalised intensity data (in relative optical density) were expressed as a percentage of vehicle and analysed via three-way ANOVA with Tukey post-hoc multiple comparisons with results considered significant if p Analysis of WFA staining intensity revealed significant main effects of BDNF (F1, 982 = 45.46, p Our data demonstrate that BDNF increased GAD65/67 levels and PNN density in developing cortical interneurons. Our data suggest this enhancing effect of BDNF on GAD65/67 and PNN maturation is dependent on JNK signalling. Given evidence of reduced BDNF levels in schizophrenia patients, we speculate that impairments in MAP2K7-JNK signalling may be linked with functional BDNF deficits in schizophrenia and that BDNF disruption could be involved in disorder aetiology and pathophysiology.

A lack of GluN2A-containing NMDA receptors confers a vulnerability to redox dysregulation: Consequences on parvalbumin interneurons, and their perineuronal nets

The GluN2A subunit of NMDA receptors (NMDARs) plays a critical role during postnatal brain development as its expression increases while Glun2B expression decreases. Mutations and polymorphisms in GRIN2A gene, coding for GluN2A, are linked to developmental brain disorders such as mental retardation, epilepsy, schizophrenia. Published data suggest that GluN2A is involved in maturation and phenotypic maintenance of parvalbumin interneurons (PVIs), and these interneurons suffer from a deficient glutamatergic neurotransmission via GluN2A-containing NMDARs in schizophrenia.In the present study, we find that although PVIs and their associated perineuronal nets (PNNs) appear normal in anterior cingulate cortex of late adolescent/young adult GRIN2A KO mice, a lack of GluN2A delays PNN maturation. GRIN2A KO mice display a susceptibility to redox dysregulation as sub-threshold oxidative stress and subtle alterations in antioxidant systems are observed in their prefrontal cortex. Consequently, an oxidative insult applied during early postnatal development increases oxidative stress, decreases the number of parvalbumin-immunoreactive cells, and weakens the PNNs in KO but not WT mice. These effects are long-lasting, but preventable by the antioxidant, N-acetylcysteine. The persisting oxidative stress, deficit in PVIs and PNNs, and reduced local high-frequency neuronal synchrony in anterior cingulate of late adolescent/young adult KO mice, which have been challenged by an early-life oxidative insult, is accompanied with microglia activation.Altogether, these indicate that a lack of GluN2A-containing NMDARs alters the fine control of redox status, leading to a delayed maturation of PNNs, and conferring vulnerability for long-term oxidative stress, microglial activation, and PVI network dysfunction.

Differential Expression and Cell-Type Specificity of Perineuronal Nets in Hippocampus, Medial Entorhinal Cortex, and Visual Cortex Examined in the Rat and Mouse.

Perineuronal nets (PNNs) are specialized extracellular matrix (ECM) structures that condense around the soma and proximal dendrites of subpopulations of neurons. Emerging evidence suggests that they are involved in regulating brain plasticity. However, the expression of PNNs varies between and within brain areas. A lack of quantitative studies describing the distribution and cell-specificity of PNNs makes it difficult to reveal the functional roles of PNNs. In the current study, we examine the distribution of PNNs and the identity of PNN-enwrapped neurons in three brain areas with different cognitive functions: the dorsal hippocampus, medial entorhinal cortex (mEC) and primary visual cortex (V1). We compared rats and mice as knowledge from these species are often intermingled. The most abundant expression of PNNs was found in the mEC and V1, while dorsal hippocampus showed strikingly low levels of PNNs, apart from dense expression in the CA2 region. In hippocampus we also found apparent species differences in expression of PNNs. While we confirm that the PNNs enwrap parvalbumin-expressing (PV+) neurons in V1, we found that they mainly colocalize with excitatory CamKII-expressing neurons in CA2. In mEC, we demonstrate that in addition to PV+ cells, the PNNs colocalize with reelin-expressing stellate cells. We also show that the maturation of PNNs in mEC coincides with the formation of grid cell pattern, while PV+ cells, unlike in other cortical areas, are present from early postnatal development. Finally, we demonstrate considerable effects on the number of PSD-95-gephyrin puncta after enzymatic removal of PNNs.

Effect of chronic stress during adolescence in prefrontal cortex structure and function

Critical periods of plasticity (CPPs) are defined by developmental intervals wherein neuronal circuits are most susceptible to environmental influences. The CPP of the prefrontal cortex (PFC), which controls executive functions, extends up to early adulthood and, like other cortical areas, reflects the maturation of perineuronal nets (PNNs) surrounding the cell bodies of specialized inhibitory interneurons. The aim of the present work was to evaluate the effect of chronic stress on both structure and function of the adolescent’s rat PFC. We subjected P28 rats to stressful situations for 7, 15 and 35days and evaluated the spatial distribution of histochemically-labeled PNNs in both the Medial Prefrontal Cortex (MPFC) and the Orbitofrontal Cortex (OFC) and PFC-associated behavior as well. Chronic stress affects PFC development, slowing PNN maturation in both the (MPFC) and (OFC) while negatively affecting functions associated with these areas. We speculate upon the risks of prolonged exposure to stressful environments in human adolescents and the possibility of stunted development of executive functions.

6.131 NEURODEVELOPMENT OF PARVALBUMIN CELLS AND PERINEURONAL NETS FOLLOWING EARLY LIFE TRAUMA

Early life trauma, particularly during the developmental sensitive period for infant attachment, is a major risk factor for later-life vulnerability to psychiatric disorders characterized by heightened fearfulness. The basolateral amygdala (BLA), long known for its integral role in threat response behavior, has recently been shown to be hyper-responsive in juveniles following early life abuse from the caregiver. However, the anatomical substrates that mediate increased responsiveness of the BLA at a synaptic level are unknown. Fast-spiking parvalbumin positive (PV+) inhibitory interneurons are known to play a major role in defining critical periods of neurodevelopment and have been shown to play a critical role in BLA fear response. A class of extracellular molecules referred to as perineuronal nets (PNNs), have been shown to end critical periods of neurodevelopment by locking axo-somatic synapses onto PV+ somata. Our objective is to establish whether early life abuse affects PV+ cell and PNN maturation, and whether dissolution of PNNs during healthy maternal care will correct abnormal threat response behavior in abused pups.