Epidermal Growth Factor Suppresses the Development of GABAergic Neurons Via the Modulation of Perineuronal Net Formation in the Neocortex of Developing Rodent Brains.

Previously, we reported that epidermal growth factor (EGF) suppresses GABAergic neuronal development in the rodent cortex. Parvalbumin-positive GABAergic neurons (PV neurons) have a unique extracellular structure, perineuronal nets (PNNs). PNNs are formed during the development of PV neurons and are mainly formed from chondroitin sulfate (CS) proteoglycans (CSPGs). We examined the effect of EGF on CSPG production and PNN formation as a potential molecular mechanism for the inhibition of inhibiting GABAergic neuronal development by EGF. In EGF-overexpressing transgenic (EGF-Tg) mice, the number of PNN-positive PV neurons was decreased in the cortex compared with that in wild-type mice, as in our previous report. The amount of CS and neurocan was also lower in the cortex of EGF-Tg mice, with a similar decrease observed in EGF-treated cultured cortical neurons. PD153035, an EGF receptor (ErbB1) kinase inhibitor, prevented those mentioned above excess EGF-induced reduction in PNN. We explored the molecular mechanism underlying the effect of EGF on PNNs using fluorescent substrates for matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinases (ADAMs). EGF increased the enzyme activity of MMPs and ADAMs in cultured neurons. These enzyme activities were also increased in the EGF-Tg mice cortex. GM6001, a broad inhibitor of MMPs and ADAMs, also blocked EGF-induced PNN reductions. Therefore, EGF/EGF receptor signals may regulate PNN formation in the developing cortex.

Schizophrenia Animal Modeling with Epidermal Growth Factor and Its Homologs: Their Connections to the Inflammatory Pathway and the Dopamine System.

Epidermal growth factor (EGF) and its homologs, such as neuregulins, bind to ErbB (Her) receptor kinases and regulate glial differentiation and dopaminergic/GABAergic maturation in the brain and are therefore implicated in schizophrenia neuropathology involving these cell abnormalities. In this review, we summarize the biological activities of the EGF family and its neuropathologic association with schizophrenia, mainly overviewing our previous model studies and the related articles. Transgenic mice as well as the rat/monkey models established by perinatal challenges of EGF or its homologs consistently exhibit various behavioral endophenotypes relevant to schizophrenia. In particular, post-pubertal elevation in baseline dopaminergic activity may illustrate the abnormal behaviors relevant to positive and negative symptoms as well as to the timing of this behavioral onset. With the given molecular interaction and transactivation of ErbB receptor kinases with Toll-like receptors (TLRs), EGF/ErbB signals are recruited by viral infection and inflammatory diseases such as COVID-19-mediated pneumonia and poxvirus-mediated fibroma and implicated in the immune-inflammatory hypothesis of schizophrenia. Finally, we also discuss the interaction of clozapine with ErbB receptor kinases as well as new antipsychotic development targeting these receptors.

The dual role of dopamine in the modulation of information processing in the prefrontal cortex underlying social behavior.

Dopamine in the prefrontal cortex is essential for the regulation of social behavior. However, stress-causing social withdrawal also promotes dopamine release in the prefrontal cortex. Thus, this evidence suggests opposite functions of dopamine in the prefrontal cortex. However, the influence of dopamine on prefrontal functions is yet to be fully understood. Here, we show that dopamine differentially modulated the neuronal activity triggered by social stimuli in the prefrontal cortex, depending on the duration of the dopamine activation (transient or sustained activation). Using chemogenetic techniques, we have found that social behavior was negatively regulated by a sustained increase in dopamine neuronal activity in the ventral tegmental area, while it was positively regulated by an acute increase. The duration of social interactions was positively correlated with the transient dopamine release triggered by social stimuli in the prefrontal cortex and negatively correlated with the sustained increase in prefrontal dopamine levels. Furthermore, the elevation of neural calcium signal, triggered by social stimuli, in the prefrontal cortex was attenuated by the persistent elevation of prefrontal dopamine levels, whereas an acute increase in dopamine levels enhanced it. Additionally, the chronic excess of dopamine suppressed c-Fos induction triggered by social stimuli in prefrontal neurons expressing dopamine D1 receptors, but not D2 receptors. These results suggest that sustained activation of prefrontal dopamine, at the opposite of its transient activation, can reduce prefrontal activity associated with social behavior, even for identical dopamine concentrations. Thus, dopamine plays opposite roles in modulating prefrontal activity depending on the duration of its action.

Neuregulin-1 signals from the periphery regulate AMPA receptor sensitivity and expression in GABAergic interneurons in developing neocortex.

Neuregulin-1 (NRG1) signaling is thought to contribute to both neuronal development and schizophrenia neuropathology. Here, we describe the developmental effects of excessive peripheral NRG1 signals on synaptic activity and AMPA receptor expression of GABAergic interneurons in postnatal rodent neocortex. A core peptide common to all NRG1 variants (eNRG1) was subcutaneously administered to mouse pups. Injected eNRG1 penetrated the blood-brain barrier and activated ErbB4 NRG1 receptors in the neocortex, in which ErbB4 mRNA is predominantly expressed by parvalbumin-positive GABAergic interneurons. We prepared neocortical slices from juvenile mice that were receiving eNRG1 subchronically and recorded inhibitory synaptic activity from layer V pyramidal neurons. Postnatal eNRG1 treatment significantly enhanced polysynaptic IPSCs, although monosynaptic IPSCs were not affected. Examination of excitatory inputs to parvalbumin-containing GABAergic interneurons revealed that eNRG1 treatment significantly increased AMPA-triggered inward currents and the amplitudes and frequencies of miniature EPSCs (mEPSCs). Similar effects on mEPSCs were observed in mice treated with a soluble, full-length form of NRG1 type I. Consistent with the electrophysiologic data, expression of the AMPA receptor GluA1 (i.e., GluR1, GluRA) was upregulated in the postsynaptic density/cytoskeletal fraction prepared from eNRG1-treated mouse neocortices. Cortical GABAergic neurons cultured with eNRG1 exhibited a significant increase in surface GluA1 immunoreactivity at putative synaptic sites on their dendrites. These results indicate that NRG1 circulating in the periphery influences postnatal development of synaptic AMPA receptor expression in cortical GABAergic interneurons and may play a role in conditions characterized by GABA-associated neuropathologic processes.

Expression of ErbB4 in substantia nigra dopamine neurons of monkeys and humans.

Abnormal neuregulin-1 signaling through its receptor (ErbB4) might be associated with schizophrenia, although their neuropathological contribution remains controversial. To assess the role of neuregulin-1 in the dopamine hypothesis of schizophrenia, we used in situ hybridization and immunoblotting to investigate the cellular distribution of ErbB4 mRNA in the substantia nigra of Japanese monkeys (Macaca fuscata) and human postmortem brains. In both monkeys and humans, significant signal for ErbB4 mRNA was detected in substantia nigra dopamine neurons, which were identified by melanin deposits. The expression of ErbB4 mRNA in nigral dopamine neurons was confirmed with an independent RNA probe, as well as with combined tyrosine hydroxylase immunostaining. Immunoblotting appeared to support the observation of in situ hybridization. Immunoreactivity for ErbB4 protein was much more enriched in substantia nigra pars compacta containing dopamine neurons than in neighboring substantia nigra pars reticulata. These observations suggest that ErbB4 is expressed in the dopaminergic neurons of primate substantia nigra and ErbB4 abnormality might contribute to the dopaminergic pathology associated with schizophrenia or other brain diseases.

Activation of epidermal growth factor receptor ErbB1 attenuates inhibitory synaptic development in mouse dentate gyrus.

Ligands for the epidermal growth factor receptor ErbB1, such as epidermal growth factor (EGF) and transforming growth factor alpha (TGFalpha), negatively regulate synaptic maturation of GABAergic neurons in the developing neocortex. Here, we evaluated the effects of these ligands in vivo on developing inhibitory neurons in the dentate gyrus. Hippocampal slices were prepared from postnatal mice repeatedly challenged with EGF or from transgenic mice overexpressing TGFalpha. We monitored paired pulse depression of field population spikes evoked by perforant path stimulation to estimate the strength of local inhibition. Administration of EGF increased the paired pulse ratio, suggesting a reduction of inhibitory strength. A similar reduction was observed in TGFalpha transgenic mice. Monitoring miniature and evoked synaptic currents, we estimated EGF effects on synaptic input and output of GABAergic neurons. EGF treatment diminished the amplitude of excitatory postsynaptic currents (EPSCs) in the GABAergic neurons without affecting their miniature EPSCs. EGF also affected output strength of the GABAergic neurons: The frequency of miniature inhibitory postsynaptic currents (IPSCs) and the evoked IPSC/evoked EPSC ratio were decreased in granule cells. In parallel, EGF down-regulated the protein level of vesicular GABA transporter. Thus, ErbB1 ligands influence GABAergic inhibitory synaptic transmission in the developing dentate gyrus.

A cyclooxygenase-2 inhibitor ameliorates behavioral impairments induced by striatal administration of epidermal growth factor.

Consistent with the hypothesis that neuroinflammatory processes contribute to the neuropathology of schizophrenia, the protein levels of epidermal growth factor (EGF) and its receptor ErbB1 are abnormal in patients with schizophrenia. To evaluate neuropathological significance of this abnormality, we established an animal model for behavioral deficits by administering EGF into the striatum and evaluated the effects of cyclooxygenase-2 (Cox-2) inhibitor celecoxib. Intracranial infusion of EGF into the striatum of adult male rats activated ErbB1 and induced neurobehavioral impairments observed in several schizophrenia models. Unilateral EGF infusion to the striatum lowered prepulse inhibition (PPI) in a dose-dependent manner and impaired latent learning of active shock avoidance without affecting basal learning ability. Bilateral EGF infusion similarly affected PPI. In contrast, EGF infusion to the nucleus accumbens did not induce a behavioral deficit. Intrastriatal EGF infusion also increased Cox-2 expression, elevated tyrosine hydroxylase activity, and upregulated the levels of dopamine and its metabolites. Subchronic administration of celecoxib (10 mg/kg, p.o.) ameliorated the abnormalities in PPI and latent learning as well as normalized dopamine metabolism. We conclude that this EGF-triggered neuroinflammatory process is mediated in part by Cox-2 activity and perturbs dopamine metabolism to generate neurobehavioral abnormalities.

Neonatal exposure to epidermal growth factor induces dopamine D2-like receptor supersensitivity in adult sensorimotor gating.

RATIONALE: Abnormality in the neurotrophic factor for dopamine neurons, epidermal growth factor (EGF), is associated with schizophrenia. Thus, rats treated with EGF as neonates are used as a putative animal model for schizophrenia showing impaired prepulse inhibition (PPI) and other cognitive deficits in the adult stage. OBJECTIVES: To elucidate the abnormal behavioral traits of this animal model, the EGF effects on the dopaminergic system were analyzed pharmacologically and biochemically at the adult stage. RESULTS: We examined the effects of subthreshold doses of dopamine agonists on PPI in this model. A non-selective dopamine agonist, apomorphine (0.1 mg/kg), decreased PPI in EGF-treated rats, but not in controls. Further, a D(2)-like receptor agonist, quinpirole (0.01 and 0.03 mg/kg), similarly decreased PPI in EGF-treated rats but had no effect in the control animals. In contrast, a D(1)-like receptor agonist, SKF38393 (3 and 10 mg/kg), had no effect on PPI in both groups. To explore the molecular mechanism underlying the change in sensorimotor gating, we assessed D(1) and D(2) receptors expression in the prefrontal cortex, striatum and hippocampus and their downstream signaling. Although there were no significant differences in basal receptor levels, quinpirole administration significantly enhanced phosphorylation of extracellular signal-regulated kinase (ERK) and cAMP response element binding protein (CREB) in the striatum of EGF-treated rats. CONCLUSION: These results suggest that circulating EGF in the early development substantially influences D(2) receptor-dependent regulation of sensorimotor gating.

Transforming growth factor alpha attenuates the functional expression of AMPA receptors in cortical GABAergic neurons.

In the developing neocortex, brain-derived neurotrophic factor (BDNF) exerts a trophic activity to increase the expression and channel activity of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)-type glutamate receptor subunits. Here, we demonstrate that the epidermal growth factor (EGF) receptor (ErbB1) ligands exert the opposite biological activity in cultured neocortical neurons. Subchronic stimulation of ErbB1 with transforming growth factor alpha (TGFalpha), EGF, or heparin-binding EGF (HB-EGF) down-regulated protein expression of the GluR1 AMPA receptor subunit in cultured neocortical neurons. In agreement, TGFalpha treatment decreased the Bmax of [3H] AMPA binding and GluR1 mRNA levels. Immunocytochemistry revealed that the decrease in GluR1 was most pronounced in multipolar GABAergic neurons. To examine the physiological consequences, we recorded AMPA-evoked currents as well as miniature excitatory postsynaptic currents in morphologically identified putative GABAergic neurons in culture. Subchronic TGFalpha treatment decreased AMPA-triggered currents as well as the amplitude and frequency of miniature excitatory postsynaptic currents. An ErbB1 tyrosine kinase inhibitor, PD153035, inhibited the TGFalpha effect. Moreover, TGFalpha counteracted the neurotrophic activity of BDNF on AMPA receptor expression. Co-application of TGFalpha with BDNF blocked the BDNF-triggered up-regulation of AMPA receptor expression and currents. These observations reveal a negative regulatory activity of the ErbB1 ligand, TGFalpha, which reduces the input sensitivity of cortical GABAergic neurons to attenuate their inhibitory function.

Brain-derived neurotrophic factor induces mammalian target of rapamycin-dependent local activation of translation machinery and protein synthesis in neuronal dendrites.

In neurons, perisynaptic or dendritic translation is implicated in synapse-wide alterations of function and morphology triggered by neural activity. The molecular mechanisms controlling local translation activation, however, have yet to be elucidated. Here, we show that local protein synthesis and translational activation in neuronal dendrites are upregulated by brain-derived neurotrophic factor (BDNF) in a rapamycin and small interfering RNA specific for mammalian target of rapamycin (mTOR)-sensitive manner. In parallel, BDNF induced the phosphorylation of tuberin and the activation of mTOR in dendrites and the synaptoneurosome fraction. mTOR activation stimulated translation initiation processes involving both eIF4E/4E-binding protein (4EBP) and p70S6 kinase/ribosomal S6 protein. BDNF induced phosphorylation of 4EBP in isolated dendrites. Moreover, local puff application of BDNF to dendrites triggered S6 phosphorylation in a restricted area. Taken together, these data indicate that mTOR-dependent translation activation is essential for the upregulation of local protein synthesis in neuronal dendrites.