Neuregulin-1-dependent control of amygdala microcircuits is critical for fear extinction.

The posttraumatic stress disorder is marked by an impaired ability to extinct fear memory acquired in trauma. Although previous studies suggest that fear extinction depends on the function of the amygdala, the underlying mechanisms are unclear. We found that NRG1 receptors (ErbB4) were abundantly expressed in the intercalated cells mass of amygdala (ITC). The NRG1-ErbB4 pathway in the ITC promotes fear extinction. The NRG1-ErbB4 pathway in the ITC did not affect excitatory input to ITC neurons from BLA neurons but increased feed-forward inhibition of (the central medial nucleus of the amygdala) CeM neurons through increased GABAergic neurotransmission of ITC neurons. We also found that the NRG1-ErbB4 signaling pathway in ITC might regulate fear extinction through P/Q-type voltage-activated Ca2+ channels (VACCs) but not through L- or N-type VACCs. Overall, our results suggest that the NRG1-ErbB4 signaling pathway in the ITC might represent a potential target for the treatment of anxiety disorders.

Roles and mechanisms of NRG1 in modulating the pathogenesis of NAFLD through ErbB3 signaling in hepatocytes (NRG1 modulates NAFLD through ErbB3 signaling).

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is an emerging chronic liver disease. However, the underlying mechanisms remained poorly understood. Neuregulin (NRG) family participate in energy metabolism, and might be related to NAFLD. METHODS: L02 cells were exposed to oleic acid to establish a cellular model of NAFLD. We analyzed the NAFLD cells with NRG1 and subsequent ErbB3 siRNA treatment. Cellular total lipid was stained by Oil Red O, while triglyceride content and inflammation markers were measured by enzymatic kits. The expressions of down-stream molecules were evaluated by western blot. RESULTS: In vitro, NRG1 could alleviate the steatosis of NAFLD, and inhibit the expression of IL-6 and TNF-α. The downregulation of ErbB3 aggravated steatosis, improved the levels of triglyceride, IL-6 and TNF-α in NRG1-treated NAFLD. Moreover, NRG1 treatment up-regulated ErbB3 phosphorylation, and increased the expression of PI3K and phosphorylation-AKT. When NRG1-treated NAFLD cells were transfected with ErbB3 siRNA, the expressions of ErbB3, p-ErbB3, p-AKT and PI3K were all reduced. CONCLUSION: NRG1 might play a protective role in the pathogenesis of NAFLD through ErbB3 phosphorylation to modulate the activation of PI3K-AKT pathway. The findings will expand the understanding of the mechanisms of NAFLD, and provide potential therapeutic targets.

Heregulin-induced cell migration is prevented by trastuzumab and trastuzumab-emtansine in HER2+ breast cancer.

PURPOSE: Heregulin (HRG) signaling has been implicated in the development of an aggressive phenotype in breast cancer (BC) cells, and HER2 overexpression has been associated with a worse prognosis in BC patients. Nevertheless, the molecular mechanisms through which HRG affects the efficiency of anti-HER2 therapies such as trastuzumab (Tz) and trastuzumab-emtansine (T-DM1) are currently unknown. METHODS: In the present study, we evaluate the molecular action of HRG toward fundamental scaffold proteins and several kinases in the signal transduction pathways triggered via HER2/HER3, which integrate precise and sequential steps to promote changes in cell morphology to impulse BC cell migration. In addition, we evaluate the effectiveness of Tz and T-DM1 on the control of key proteins involved in BC cell motility, since the acquisition of a migratory phenotype is essential to promote invasion and metastasis. RESULTS: We show that HRG induces actin cytoskeleton reorganization and focal adhesion complex formation, and promotes actin nucleation in BT-474 BC cells. This signaling is triggered by HER2/HER3 to c-Src, FAK and paxillin. When paxillin is phosphorylated, it recruits PAK1, which then phosphorylates cortactin. In parallel, paxillin signals to N-WASP, and both signalings regulate Arp2/3 complex, leading to the local reorganization of actin fibers. CONCLUSIONS: Our findings reveal an original mechanism by which HRG increases HER2+ BC cell motility, and show that the latter can be abolished by Tz and T-DM1 treatments. These results provide evidence for the molecular mechanisms involved in cell motility and drug resistance. They will be useful to develop new and more specific therapeutic schemes that interfere with the progression and metastasis of HER2+ BC.

Behavioural effects of high fat diet in adult Nrg1 type III transgenic mice.

Some diets appear to have detrimental effects on schizophrenia symptoms. Neuregulin 1 (NRG1) is a risk gene for schizophrenia and a recently developed transgenic mouse model for Nrg1 type III demonstrates a schizophrenia-relevant phenotype. The current study evaluated the behavioural response of Nrg1 type III transgenic mice to a high fat diet (HFD) to determine the potential interactive impact of diets and genetic risk factors on disease symptoms. Male and female Nrg1 III and control littermates (N = 13-24) were exposed during adulthood to either HFD or standard chow diet (CHOW) for eight weeks before being tested in behavioural domains relevant to schizophrenia. Locomotion and exploration, anxiety, social behaviours (including social preference), sensorimotor gating (i.e. prepulse inhibition, PPI), associative learning, and anhedonia were assessed. HFD increased the body weight gain of mice, suppressed locomotion, exploration, and anxiety-related behaviours in a sex-dependent manner. HFD augmented the PPI response in male mice and decreased anhedonia in a sucrose preference test. Finally, HFD had a sex-dependent impact on fear-associated memory with HFD-induced cognitive impairments being most prominent in Nrg1 transgenic females. In conclusion, HFD and mutant Nrg1 III interactively impair particular cognitive domains in a sex-specific manner. Thus, our preclinical data suggest that genetic predisposition to the schizophrenia risk gene NRG1 may modulate detrimental behavioural effects of diets. This indicates the importance to research further the role of particular diets in the context of populations at risk to develop schizophrenia.

Neuregulin-1-ErbB signaling promotes microglia activation contributing to mechanical allodynia of cyclophosphamide-induced cystitis.

AIMS: Central sensitization playsimportant roles in cyclophosphamide (CYP)-induced cystitis. In addition, as a visceral pain, CYP-induced chronic pain shares common pathophysiological mechanisms with neuropathic pain. Previous studies demonstrated that neuregulin-1 (Nrg1)-ErbB signaling contributes to neuropathic pain, but whether and how this signaling influences mechanical allodynia in CYP-induced cystitis is unclear. This study aimed to determine whether and how Nrg1-ErbB signaling modulates mechanical allodynia in a CYP-induced cystitis rat model. METHODS: Systemic injection with CYP was used to establish a rat model of bladder pain syndrome/interstitial cystitis (BPS/IC). An irreversible ErbB family receptor inhibitor, PD168393, and exogenous Nrg1 were intrathecally injected to modulate Nrg1-ErbB signaling. Mechanical allodynia in the lower abdomen was assessed with von-Frey filaments using the up-down method. Western blot analysis and immunofluorescence staining were used to measure the expression of Nrg1-ErbB signaling, Iba-1, p-p38, and IL-1ß in the L6-S1 spinal dorsal horn (SDH). RESULTS: We observed upregulation of Nrg1-ErbB signaling as well as overexpression of the microglia activation markers Iba-1 and p-p38 and the proinflammatory factor, interleukin-1ß (IL-1ß), in the SDH of the cystitis group. Further, treatment with PD168393 attenuated mechanical allodynia in CYP-induced cystitis and inhibited microglia activation, leading to decreased production of IL-1ß. The inhibitor PD168393 reversed the algesic effect of exogenous Nrg1 on the cystitis model. CONCLUSIONS: Nrg1-ErbB signaling may promote microglia activation, contributing to mechanical allodynia of CYP-induced cystitis. Our study showed that modulation of Nrg1-ErbB signaling may have therapeutic value for treating pain symptoms in BPS/IC.

Sarcopenia in Cirrhosis: Looking Beyond the Skeletal Muscle Loss to See the Systemic Disease.

Sarcopenia is a common complication of cirrhosis and is defined as a progressive and generalized loss of skeletal muscle mass, strength, and function. Sarcopenia is associated with poor prognosis and increased mortality. How sarcopenia and muscle wasting relate to such poor outcomes requires looking beyond the overt muscle loss and at this entity as a systemic disease that affects muscles of vital organs including cardiac and respiratory muscles. This review explores the pathophysiological pathways and mechanisms that culminate in poor outcomes associated with sarcopenia. This provides a launching pad to identify potential targets for therapeutic intervention and optimization to improve patient outcomes.

Localization and dynamic changes of neuregulin-1 at C-type synaptic boutons in association with motor neuron injury and repair.

C-type synaptic boutons (C-boutons) provide cholinergic afferent input to spinal cord motor neurons (MNs), which display an endoplasmic reticulum (ER)-related subsurface cistern (SSC) adjacent to their postsynaptic membrane. A constellation of postsynaptic proteins is clustered at C-boutons, including M2 muscarinic receptors, potassium channels, and σ-1 receptors. In addition, we previously found that neuregulin (NRG)1 is associated with C-boutons at postsynaptic SSCs, whereas its ErbB receptors are located in the presynaptic compartment. C-bouton-mediated regulation of MN excitability has been implicated in MN disease, but NRG1-mediated functions and the impact of various pathologic conditions on C-bouton integrity have not been studied in detail. Here, we investigated changes in C-boutons after electrical stimulation, pharmacological treatment, and peripheral nerve axotomy. SSC-linked NRG1 clusters were severely disrupted in acutely stressed MNs and after tunicamycin-induced ER stress. In axotomized MNs, C-bouton loss occurred in concomitance with microglial recruitment and was prevented by the ER stress inhibitor salubrinal. Activated microglia displayed a positive chemotaxis to C-boutons. Analysis of transgenic mice overexpressing NRG1 type I and type III isoforms in MNs indicated that NRG1 type III acts as an organizer of SSC-like structures, whereas NRG1 type I promotes synaptogenesis of presynaptic cholinergic terminals. Moreover, MN-derived NRG1 signals may regulate the activity of perineuronal microglial cells. Together, these data provide new insights into the molecular and cellular pathology of C-boutons in MN injury and suggest that distinct NRG1 isoform-mediated signaling functions regulate the complex matching between pre- and postsynaptic C-bouton elements.-Salvany, S., Casanovas, A., Tarabal, O., Piedrafita, L., Hernández, S., Santafé, M., Soto-Bernardini, M. C., Calderó, J., Schwab, M. H., Esquerda, J. E. Localization and dynamic changes of neuregulin-1 at C-type synaptic boutons in association with motor neuron injury and repair.

Neuregulin 1 type III improves peripheral nerve myelination in a mouse model of congenital hypomyelinating neuropathy.

Myelin sheath thickness is precisely regulated and essential for rapid propagation of action potentials along myelinated axons. In the peripheral nervous system, extrinsic signals from the axonal protein neuregulin 1 (NRG1) type III regulate Schwann cell fate and myelination. Here we ask if modulating NRG1 type III levels in neurons would restore myelination in a model of congenital hypomyelinating neuropathy (CHN). Using a mouse model of CHN, we improved the myelination defects by early overexpression of NRG1 type III. Surprisingly, the improvement was independent from the upregulation of Egr2 or essential myelin genes. Rather, we observed the activation of MAPK/ERK and other myelin genes such as peripheral myelin protein 2 and oligodendrocyte myelin glycoprotein. We also confirmed that the permanent activation of MAPK/ERK in Schwann cells has detrimental effects on myelination. Our findings demonstrate that the modulation of axon-to-glial NRG1 type III signaling has beneficial effects and improves myelination defects during development in a model of CHN.

NRG1 functions downstream of EDS1 to regulate TIR-NLR-mediated plant immunity in Nicotiana benthamiana.

Effector-triggered immunity (ETI) in plants involves a large family of nucleotide-binding leucine-rich repeat (NLR) immune receptors, including Toll/IL-1 receptor-NLRs (TNLs) and coiled-coil NLRs (CNLs). Although various NLR immune receptors are known, a mechanistic understanding of NLR function in ETI remains unclear. The TNL Recognition of XopQ 1 (Roq1) recognizes the effectors XopQ and HopQ1 from Xanthomonas and Pseudomonas, respectively, which activates resistance to Xanthomonas euvesicatoria and Xanthomonas gardneri in an Enhanced Disease Susceptibility 1 (EDS1)-dependent way in Nicotiana benthamiana In this study, we found that the N. benthamiana N requirement gene 1 (NRG1), a CNL protein required for the tobacco TNL protein N-mediated resistance to tobacco mosaic virus, is also essential for immune signaling [including hypersensitive response (HR)] triggered by the TNLs Roq1 and Recognition of Peronospora parasitica 1 (RPP1), but not by the CNLs Bs2 and Rps2, suggesting that NRG1 may be a conserved key component in TNL signaling pathways. Besides EDS1, Roq1 and NRG1 are necessary for resistance to Xanthomonas and Pseudomonas in N. benthamiana NRG1 functions downstream of Roq1 and EDS1 and physically associates with EDS1 in mediating XopQ-Roq1-triggered immunity. Moreover, RNA sequencing analysis showed that XopQ-triggered gene-expression profile changes in N. benthamiana were almost entirely mediated by Roq1 and EDS1 and were largely regulated by NRG1. Overall, our study demonstrates that NRG1 is a key component that acts downstream of EDS1 to mediate various TNL signaling pathways, including Roq1 and RPP1-mediated HR, resistance to Xanthomonas and Pseudomonas, and XopQ-regulated transcriptional changes in N. benthamiana.

Neuregulin-1ß modulates myogenesis in septic mouse serum-treated C2C12 myotubes in vitro through PPARγ/NF-κB signaling.

Sepsis-induced skeletal muscle atrophy is a pathological condition characterized by the loss of strength and muscle mass. Cytokine-induced apoptosis and impaired myogenesis play key roles in the development of this condition. However, the complete underlying mechanism remains largely unknown. Neuregulins are glial growth factors essential for myogenesis that regulate muscle metabolism. We investigated the role of neuregulin-1ß (NRG-1ß) in sepsis-induced apoptosis and myogenesis in skeletal muscle using a serum-based in vitro sepsis model. C2C12 myoblasts were differentiated by treatment with proliferative medium for 7 days. Then, cells were treated with 2% sham mouse serum, 1 nM NRG-1ß in 2% sham mouse serum, 2% septic mouse serum (SMS), or 1 nM NRG-1ß in 2% SMS. Exposure to SMS induced apoptosis, impaired myogenesis, and downregulated PPARγ. NRG-1ß co-incubation remedied all these effects and inhibited NF-κB transcriptional activity. A specific PPARγ antagonist (GW9662) was also administered as a 2-h pretreatment to block PPARγ-mediated signaling and appeared to attenuate the effects of NRG-1ß. Taken together, our results demonstrate that NRG-1ß functions via a PPARγ/NF-κB-dependent pathway to modulate myogenesis and protect against apoptosis in SMS-treated C2C12 myotubes.

ß-Heregulin impairs EGF induced PLC-γ1 signalling in human breast cancer cells.

The interplay of ErbB receptor homo- and heterodimers plays a crucial role in the pathology of breast cancer since activated signal transduction cascades coordinate proliferation, survival and migration of cells. EGF and ß-Heregulin are well characterised ligands known to induce ErbB homo- and heterodimerisation, which have been associated with disease progression. In the present study, we investigated the impact of both factors on the migration of MDA-NEO and MDA-HER2 human breast cancer cells. MDA-NEO cells are positive for EGFR and HER3, while MDA-HER2 cells express EGFR, HER2 and HER3. Cell migration analysis revealed that ß-Heregulin potently impaired EGF induced migration in both cell lines. Western blot studies showed that both ErbB receptor and PLC-γ1 tyrosine phosphorylation levels were diminished in EGF and ß-Heregulin co-treated MDA-NEO and MDA-HER2 cells, which was further correlated to a significantly impaired calcium influx. Our data indicate that EGF and HRG may interfere with each other for receptor binding and dimerisation, which ultimately has an impact on signalling outcome.

Evaluation of neuregulin-1's neuroprotection against ischemic injury in rats using diffusion tensor imaging.

Stroke is a devastating neurovascular disorder that results in damage to neurons and white matter tracts. It has been previously demonstrated that neuregulin-1 (NRG-1) protects neurons from ischemic injury following stroke. Here, diffusion tensor imaging (DTI) was utilized to characterize the effects of NRG-1 treatment on cererbral infarction and integrity of white matter after ischemic insult using a permanent middle celebral artery occlusion (pMCAo) rat model. In the present study, sixteen Sprague-Dawley rats underwent pMCAo surgery and received either a single intra-arterial bolus (20 µg/kg) dose of NRG-1 or saline immediately prior to pMCAo. MRI including T2-weighted imaging and DTI was performed in the first 3 h post stroke, and repeated 48 h later. It is found that the stroke infarction was significantly reduced in the NRG-1 treated group. Also, NRG-1 prevented the reduction of fractional anisotropy (FA) in white matter tracts of fornix and corpus callosum (CC), indicating its protection of CC and fornix white matter bundles from ischemia insult. As a conclusion, the present DTI results demonstrate that NRG-1 has significantly neuroprotective effects in both cerebral cortex and white matter including corpus callosum and fornix during acute stroke. In particular, NRG-1 is more effective on stroke lesion with mild ischemia. As CC and fornix white matter bundles play critical roles in transcallosal connectivity and hippocampal projections respectively in the central nervous system, the findings could provide complementary information for better understanding the biological mechanism of NRG-1's neuroprotection in ischemic tissues and neurobehavioral effects.

Electrical Stimulation with a Conductive Polymer Promotes Neurite Outgrowth and Synaptogenesis in Primary Cortical Neurons in 3D.

Deficits in neurite outgrowth and synaptogenesis have been recognized as an underlying developmental aetiology of psychosis. Electrical stimulation promotes neuronal induction including neurite outgrowth and branching. However, the effect of electrical stimulation using 3D electrodes on neurite outgrowth and synaptogenesis has not been explored. This study examined the effect of 3D electrical stimulation on 3D primary cortical neuronal cultures. 3D electrical stimulation improved neurite outgrowth in 3D neuronal cultures from both wild-type and NRG1-knockout (NRG1-KO) mice. The expression of synaptophysin and PSD95 were elevated under 3D electrical stimulation. Interestingly, 3D electrical stimulation also improved neural cell aggregation as well as the expression of PSA-NCAM. Our findings suggest that the 3D electrical stimulation system can rescue neurite outgrowth deficits in a 3D culturing environment, one that more closely resembles the in vivo biological system compared to more traditionally used 2D cell culture, including the observation of cell aggregates as well as the upregulated PSA-NCAM protein and transcript expression. This study provides a new concept for a possible diagnostic platform for neurite deficits in neurodevelopmental diseases, as well as a viable platform to test treatment options (such as drug delivery) in combination with electrical stimulation.

Neuregulin 1 regulates amyloid precursor protein cell surface expression and non-amyloidogenic processing.

The amyloid precursor protein (APP) is a key molecule in Alzheimer's disease. The prevailing view is that APP is initially transported to the plasma membrane as a full-length protein. Its localization at the cell surface can trigger downstream signaling and APP cleavage. Our previous work has shown that Neuregulin 1 (NRG1) has neuroprotective effects in an Alzheimer's disease model. In the present study, we examine whether NRG1 signaling is involved in APP expression and non-amyloidogenic processing in neuronal cells. Here we show that NRG1 increased the cell surface expression of APP without changing the total amount of APP mRNA or protein expression in SH-SY5Y cells and in rat primary cortical neurons. In addition, NRG1 significantly increased the levels of the secreted form of APP, sAPPα, in the conditioned media but did not change the expression of ADAM10 on the cell surface or in the cell lysates. Furthermore, we found that the protein level of NRG1 was reduced in the hippocampus of Alzheimer's disease (AD) patients. Our results demonstrate that NRG1 increased APP expression on the cell surface and sAPPα secretion into the media of neuronal cell cultures. Taken together, these results suggest a role for NRG1 in non-amyloidogenic processing.

Reduced type III neuregulin 1 expression does not modulate the behavioural sensitivity of mice to acute Δ9-tetrahydrocannabinol (D9-THC).

Mice with a mutation in the transmembrane domain of the schizophrenia risk gene, neuregulin 1 (Nrg1 TM HET), are more susceptible to the neuro-behavioural effects of Δ9-tetrahydrocannabinol (D9-THC), the principal psychoactive component in cannabis. However, NRG1 is transcriptionally complex with over 30 different isoforms, most of which carry a transmembrane domain, raising the question which NRG1 isoform(s) may contribute to this phenotype. Type III NRG1/Nrg1 is the most brain abundant isoform and brain studies have identified increased levels of type III NRG1 mRNA in humans carrying a NRG1 risk haplotype for schizophrenia. To investigate whether mice heterozygous for type III Nrg1 (i.e. knockout: type III Nrg1+/-) are more susceptible to the behavioural effects of acute doses of D9-THC, we injected male mice with either vehicle or D9-THC (3 or 10 mg/kg) before testing them for locomotion, anxiety, social interaction, and sensorimotor gating. Acute D9-THC led to reduced locomotion and reduced social interaction, but increased anxiety in mice. Furthermore, type III Nrg1 males displayed a robust deficit in sensorimotor gating and demonstrated reduced following during social interaction across drug conditions. However, they did not show a change in behavioural susceptibility to acute D9-THC compared to controls. These results reinforce the validity of type III Nrg1+/- mice for schizophrenia research and suggest that loss of function of type III Nrg1 may not be responsible for the exaggerated response to acute D9-THC observed in heterozygous Nrg1 TM mice. This highlights the importance of careful consideration of Nrg1 isoform type differences.

Neuregulin-1 is a chemoattractant and chemokinetic molecule for trunk neural crest cells.

BACKGROUND: Trunk neural crest cells migrate rapidly along characteristic pathways within the developing vertebrate embryo. Proper trunk neural crest cell migration is necessary for the morphogenesis of much of the peripheral nervous system, melanocytes, and the adrenal medulla. Numerous molecules help guide trunk neural crest cell migration throughout the early embryo. RESULTS: The trophic factor NRG1 is a chemoattractant through in vitro chemotaxis assays and in vivo silencing via a DN-erbB receptor. Interestingly, we also observed changes in migratory responses consistent with a chemokinetic effect of NRG1 in trunk neural crest velocity. CONCLUSIONS: NRG1 is a trunk neural crest cell chemoattractant and chemokinetic molecule. Developmental Dynamics 247:888-902, 2018.. © 2018 Wiley Periodicals, Inc.

Neuregulin 1/ErbB signalling modulates hippocampal mGluRI-dependent LTD and object recognition memory.

The neurotrophic factors neuregulins (NRGs) and their receptors, ErbB tyrosine kinases, regulate neurotransmission, synaptic plasticity and cognitive functions and their alterations have been associated to different neuropsychiatric disorders. Group 1 metabotropic glutamate receptors (mGluRI)-dependent mechanisms are also altered in animal models of neuropsychiatric diseases, especially mGluRI-induced glutamatergic long-term depression (mGluRI-LTD), a form of synaptic plasticity critically involved in learning and memory. Despite this evidence, a potential link between NRGs/ErbB signalling and mGluRI-LTD has never been considered. Here, we aimed to test the hypothesis that NRGs/ErbB signalling regulates mGluRI functions in the hippocampus, thus controlling CA1 pyramidal neurons excitability and synaptic plasticity as well as mGluRI-dependent behaviors. We investigated the functional interaction between NRG1/ErbB signalling and mGluRI in hippocampal CA1 pyramidal neurons, by analyzing the effect of a pharmacological modulation of NRG1/ErbB signalling on the excitation of pyramidal neurons and on the LTD at CA3-CA1 synapses induced by an mGluRI agonist. Furthermore, we verified the involvement of ErbB signalling in mGluRI-dependent learning processes, by evaluating the consequence of an intrahippocampal in vivo injection of a pan-ErbB inhibitor in the object recognition test in mice, a learning task dependent on hippocampal mGluRI. We found that NRG1 potentiates mGluRI-dependent functions on pyramidal neurons excitability and synaptic plasticity at CA3-CA1 synapses. Further, endogenous ErbB signalling per se regulates, through mGluRI, neuronal excitability and LTD in CA1 pyramidal neurons, since ErbB inhibition reduces mGluRI-induced neuronal excitation and mGluRI-LTD. In vivo intrahippocampal injection of the ErbB inhibitor, PD158780, impairs mGluRI-LTD at CA3-CA1 synapses and affects the exploratory behavior in the object recognition test. Thus, our results identify a key role for NRG1/ErbB signalling in the regulation of hippocampal mGluRI-dependent synaptic and cognitive functions, whose alteration might contribute to the pathogenesis of different brain diseases.

Neuregulin 3 and its roles in schizophrenia risk and presentation.

Neuregulins, a four-member family of epidermal growth factor-like signaling molecules, have been studied for over two decades. They were first implicated in schizophrenia in 2002 with the detection of linkage and association at the NRG1 locus followed after a few years by NRG3. However, the associations with disease have not been very consistently observed. In contrast, association of NGR3 variants with disease presentation, specifically the presence of delusions, has been more consistent. This appears to be mediated by quantitative changes in the alternative splicing of the gene, which has also been consistently observed. Additional diseases and phenotypes, psychiatric or not, have also been connected with NRG3. These results demonstrate two important aspects of behavioral genetics research. The first is that if we only consider simple risk and fail to examine the details of each patient's individual phenotype, we will miss important insights on the disease biology. This is an important aspect of the goals of precision medicine. The second is that the functional consequences of variants are often more complex than simple alterations in levels of transcription of a particular gene, including, among others, regulation of alternative splicing. To accurately model and understand the biological consequences of phenotype-associated genetic variants, we need to study the biological consequences of each specific variant. Simply studying the consequences of a null allele of the orthologous gene in a model system, runs the risk of missing the many nuances of hypomorphic and/or gain of function variants in the genome of interest.

Neuregulin1 fine-tunes pre-, post-, and perisynaptic neuromuscular junction development.

BACKGROUND: Neuromuscular junction (NMJ) development is a multistep process mediated by coordinated interactions between the nerve terminal, target muscle, and perisynaptic Schwann cell that require constant back-and-forth communication. Retrograde and anterograde growth and differentiation factors have been postulated to participate in this communication. While neuregulin1 (NRG1) has been shown to be potent anterograde signal that activates acetylcholine receptor (AChR) transcription and clustering in vitro, its roles in NMJ development in vivo remain elusive. RESULTS: Using the model of chicken embryo, we measured the effects of NRG1 signaling during NMJ development in ovo using quantitative, sequential measures of AChR cluster size and density, pre- and postsynaptic apposition, and the alignment of perisynaptic Schwann cells. Using in ovo electroporation at early stages and a targeted soluble neuregulin antagonist through all developmental stages, we found soluble NRG1 regulates AChR cluster density and size at the earliest stage prior to nerve-AChR cluster contact. Once the nerve contacts with muscle AChRs, NRG1 has pronounced effects on presynaptic specialization and on the alignment of perisynaptic Schwann cells at endplates. CONCLUSION: These findings suggest that, while NRG1 may not be critical for overall development, it appears to be important in fine-tuning pre-, post-, and perisynaptic development of the NMJ. Developmental Dynamics 246:368-380, 2017. © 2017 Wiley Periodicals, Inc.

Irreversible changes occurring in long-term denervated Schwann cells affect delayed nerve repair.

OBJECTIVE Multiple factors may affect functional recovery after peripheral nerve injury, among them the lesion site and the interval between the injury and the surgical repair. When the nerve segment distal to the lesion site undergoes chronic degeneration, the ensuing regeneration (when allowed) is often poor. The aims of the current study were as follows: 1) to examine the expression changes of the neuregulin 1/ErbB system during long-term nerve degeneration; and 2) to investigate whether a chronically denervated distal nerve stump can sustain nerve regeneration of freshly axotomized axons. METHODS This study used a rat surgical model of delayed nerve repair consisting of a cross suture between the chronically degenerated median nerve distal stump and the freshly axotomized ulnar proximal stump. Before the suture, a segment of long-term degenerated median nerve stump was harvested for analysis. Functional, morphological, morphometric, and biomolecular analyses were performed. RESULTS The results showed that neuregulin 1 is highly downregulated after chronic degeneration, as well as some Schwann cell markers, demonstrating that these cells undergo atrophy, which was also confirmed by ultrastructural analysis. After delayed nerve repair, it was observed that chronic degeneration of the distal nerve stump compromises nerve regeneration in terms of functional recovery, as well as the number and size of regenerated myelinated fibers. Moreover, neuregulin 1 is still downregulated after delayed regeneration. CONCLUSIONS The poor outcome after delayed nerve regeneration might be explained by Schwann cell impairment and the consequent ineffective support for nerve regeneration. Understanding the molecular and biological changes occurring both in the chronically degenerating nerve and in the delayed nerve repair may be useful to the development of new strategies to promote nerve regeneration. The results suggest that neuregulin 1 has an important role in Schwann cell activity after denervation, indicating that its manipulation might be a good strategy for improving outcome after delayed nerve repair.