Dendritic Overgrowth Research Articles

Disruption of mTORC1 rescues neuronal overgrowth and synapse function dysregulated by Pten loss.

Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a negative regulator of AKT/mTOR signaling pathway. Mutations in PTEN are found in patients with autism, epilepsy, or macrocephaly. In mouse models, Pten loss results in neuronal hypertrophy, hyperexcitability, seizures, and ASD-like behaviors. The underlying molecular mechanisms of these phenotypes are not well delineated. We determined which of the Pten loss-driven aberrations in neuronal form and function are orchestrated by downstream mTOR complex 1 (mTORC1). Rapamycin-mediated inhibition of mTORC1 prevented increase in soma size, migration, spine density, and dendritic overgrowth in Pten knockout dentate gyrus granule neurons. Genetic knockout of Raptor to disrupt mTORC1 complex formation blocked Pten loss-mediated neuronal hypertrophy. Electrophysiological recordings revealed that genetic disruption of mTORC1 rescued Pten loss-mediated increase in excitatory synaptic transmission. We have identified an essential role for mTORC1 in orchestrating Pten loss-driven neuronal hypertrophy and synapse formation.

PTEN Regulates Dendritic Arborization by Decreasing Microtubule Polymerization Rate.

Phosphatase and tensin homolog (PTEN) is a major negative regulator of the phosphatidylinositol-3-kinase (PI3K)/Akt/mechanistic target of rapamycin (mTOR) pathway. Loss-of-function mutations in PTEN have been found in a subset of patients with macrocephaly and autism spectrum disorder (ASD). PTEN loss in neurons leads to somal hypertrophy, aberrant migration, dendritic overgrowth, increased spine density, and hyperactivity of neuronal circuits. These neuronal overgrowth phenotypes are present on Pten knock-out (KO) and reconstitution with autism-associated point mutations. The mechanism underlying dendritic overgrowth in Pten deficient neurons is unclear. In this study, we examined how Pten loss impacts microtubule (MT) dynamics in both sexes using retroviral infection and transfection strategies to manipulate PTEN expression and tag the plus-end MT binding protein, end-binding protein 3 (EB3). We found Pten KO neurons sprout more new processes over time compared with wild-type (WT) neurons. We also found an increase in MT polymerization rate in Pten KO dendritic growth cones. Reducing MT polymerization rate to the WT level was sufficient to reduce dendritic overgrowth in Pten KO neurons in vitro and in vivo Finally, we found that rescue of dendritic overgrowth via inhibition of MT polymerization was sufficient to improve the performance of Pten KO mice in a spatial memory task. Taken together, our data suggests that one factor underlying PTEN loss dependent dendritic overgrowth is increased MT polymerization. This opens the possibility for an intersectional approach targeting MT polymerization and mTOR with low doses of inhibitors to achieve therapeutic gains with minimal side effects in pathologies associated with loss of neuronal PTEN function.SIGNIFICANCE STATEMENT Loss of Pten function because of genetic deletion or expression of mutations associated with autism spectrum disorder (ASD), results in overgrowth of neurons including increased total dendritic length and branching. We have discovered that this overgrowth is accompanied by increased rate of microtubule (MT) polymerization. The increased polymerization rate is insensitive to acute inhibition of mechanistic target of rapamycin (mTOR)C1 or protein synthesis. Direct pharmacological inhibition of MT polymerization can slow the polymerization rate in Pten knock-out (KO) neurons to rates seen in wild-type (WT) neurons. Correction of the MT polymerization rate rescues increased total dendritic arborization and spatial memory. Our studies suggest that phosphatase and tensin homolog (PTEN) inhibits dendritic growth through parallel regulation of protein synthesis and cytoskeletal polymerization.

Coarsening and dendritic instability of spheroidal graphite in high silicon cast iron under thermal cycling in the ferritic domain

High silicon ferritic spheroidal graphite cast irons have been developed for high temperature service, in particular under thermal cycling conditions. The theoretical maximum service temperature is defined as the upper limit of the two-phase ferrite+graphite domain, which increases with the alloy silicon content. While isothermal heat treatment close to this temperature showed little evolution of the graphite distribution, thermal cycling led to a significant coarsening of the graphite particles associated with dendritic overgrowth of the large graphite particles. This unexpected behaviour is here observed and described for the first time.

FGF-FGFR Mediates the Activity-Dependent Dendritogenesis of Layer IV Neurons during Barrel Formation.

Fibroblast growth factors (FGFs) and FGF receptors (FGFRs) are known for their potent effects on cell proliferation/differentiation and cortical patterning in the developing brain. However, little is known regarding the roles of FGFs/FGFRs in cortical circuit formation. Here we show that Fgfr1/2/3 and Fgf7/9/10/22 mRNAs are expressed in the developing primary somatosensory (S1) barrel cortex. Barrel cortex layer IV spiny stellate cells (bSCs) are the primary recipients of ascending sensory information via thalamocortical axons (TCAs). Detail quantification revealed distinctive phases for bSC dendritogenesis: orienting dendrites toward TCAs, adding de novo dendritic segments, and elongating dendritic length, while maintaining dendritic patterns. Deleting Fgfr1/2/3 in bSCs had minimal impact on dendritic polarity but transiently increased the number of dendritic segments. However, 6 d later, FGFR1/2/3 loss of function reduced dendritic branch numbers. These data suggest that FGFs/FGFRs have a role in stabilizing dendritic patterning. Depolarization of cultured mouse cortical neurons upregulated the levels of several Fgf/Fgfr mRNAs within 2 h. In vivo, within 6 h of systemic kainic acid administration at postnatal day 6, mRNA levels of Fgf9, Fgf10, Fgfr2c, and Fgfr3b in S1 cortices were enhanced, and this was accompanied by exuberant dendritogenesis of bSCs by 24 h. Deleting Fgfr1/2/3 abolished kainic acid-induced bSC dendritic overgrowth. Finally, FGF9/10 gain of function also resulted in extensive dendritogenesis. Together, our data suggest that FGFs/FGFRs can be regulated by glutamate transmission to modulate/stabilize bSC dendritic complexity. Both male and female mice were used for our study.SIGNIFICANCE STATEMENT Glutamatergic transmission plays critical roles in cortical circuit formation. Its dysregulation has been proposed as a core factor in the etiology of many neurological diseases. We found that excessive glutamate transmission upregulated mRNA expression of Fgfrs and their ligands Fgfs Deleting Fgfr1/2/3 not only impaired bSC dendritogenesis but also abolished glutamate transmission-induced dendritic overgrowth. Overexpressing FGF9 or FGF10 in cortical glutamatergic neurons results in excessive dendritic outgrowth within 24 h, resembling the changes induced by excessive glutamate transmission. Our findings provide strong evidence for the physiological role of fibroblast growth factors (FGFs) and FGF receptors (FGFRs) in establishing and maintaining cortical circuits. Perturbing the expression levels of FGFs/FGFRs by excessive glutamatergic neurotransmission could lead to abnormal neuronal circuits, which may contribute to neurological and psychiatric disease.

Dendritic overgrowth and elevated ERK signaling during neonatal development in a mouse model of autism.

Autism spectrum disorder (hereafter referred to as “ASD”) is a heterogeneous neurodevelopmental condition characterized by impaired social communication and interactions, and restricted, repetitive activities or interests. Alterations in network connectivity and memory function are frequently observed in autism patients, often involving the hippocampus. However, specific changes during early brain development leading to disrupted functioning remain largely unclear. Here, we investigated the development of dendritic arbor of hippocampal CA1 pyramidal neurons in the BTBR T+tf/J (BTBR) mouse model of autism. BTBR mice display the defining behavioural features of autism, and also exhibit impaired learning and memory. We found that compared to control C57BL/6J (B6) animals, the lengths of both apical and basal dendrites were significantly greater in neonatal BTBR animals. Further, basal dendrites in the BTBR mice had higher branching complexity. In contrast, cross-sectional area of the soma was unchanged. In addition, we observed a similar density of CA1 pyramidal neurons and thickness of the neuronal layer between the two strains. Thus, there was a specific, compartmentalized overgrowth of dendrites during early development in the BTBR animals. Biochemical analysis further showed that the extracellular signal-regulated kinases (ERK) pathway was up-regulated in the hippocampus of neonatal BTBR animals. Since dendritic structure is critical for information integration and relay, our data suggest that altered development of dendrites could potentially contribute to impaired hippocampal function and behavior observed in the BTBR model, and that this might be related to increased activation of the ERK pathway.

The Etiological Role of Microglia in Autism Spectrum Disorder: A Possible Route for Early Intervention

Autism Spectrum Disorder (ASD) is a dreaded diagnosis. The treatment options are sparse and our knowledge of its etiology is woefully lacking. The purpose of this review is to outline a unifying pathway for the pathogenesis of Autism Spectrum Disorder and to describe how this sequence could be exploited to offer early intervention to patients at high risk of developing ASD. Specifically, we will describe how gestational insults can alter the lifelong functioning of fetal microglia. Those aberrant microglia are unable to properly fulfill their roles in cortical differentiation as well as synaptogenesis and the resulting cortical disorganization plus dendritic overgrowth may be responsible for the characteristic Autistic phenotype. This pathogenic pathway presents several opportunities for intervention. If applied early enough, it is possible that these therapies could alleviate some of the symptoms of ASD.

THE CORRELATION BETWEEN SMALL INTESTINAL BACTERIA OVERGROWTH AND DENDRITIC CELLS PHENOTYPE AND FUNCTION IN CIRRHOTIC PATIENTS WITH HEPATITIS B

Objective: To investigate the relationship between dendritic cells (DCs) and small intestinal bacteria overgrowth (SIBO) in cirrhotic patients with hepatitis B. Methods: Flow cytometry was used to analyze the phenotype of peripheral blood DCs in 47 cirrhotic patients with hepatitis B and 15 healthy controls. Small intestinal bacterial overgrowth was determined by lactulose hydrogen breath test (LHBT). A basal breath hydrogen >20 ppm or arise by >12 ppm above baseline following lactulose administration was taken as positive test. LHBT aggregate value, as another indicator to evaluate the growth of the small intestinal bacteria, is the sum of the LHBT within 90 min. Plasma total bilirubin and plasma albumin were measured in all subjects. Data analysis was using SPSS 17.0. Results: Of the 47 cirrhotics, 22 (46.8%) had SIBO, compared to healthy controls (6.7%; P= 0.005).The prevalence of SIBO increased with Child Pugh classification of cirrhosis (A18.2%, B 43.8% and C65.0%). The prevalence of SIBO in cirrhotic patients of Child Pugh class B and C was significantly higher than that of healthy controls (P=0.037; P=0.001). While the prevalence of SIBO in cirrhotic patients of Child Pugh class A was no statistically significant than that of healthy controls (P=0.556). The prevalence of SIBO in cirrhotic patients of de compensatory phase was significantly higher than that of compensatory phase (χ2 = 4.727, P=0.030). The plasma total bilirubin was positively correlated with LHBT aggregate value (r=0.907, P=0.000), While plasma albumin was negatively correlated (Ir= 0.810 P=0.000). The expressions of CD1a, CD80, CD83 on the surface of DCs in cirrhotic patients were all significant lower than that in healthy controls (P<0.05), while the expression of HLA DR was no statistically significant than that in healthy control. The expressions rate of CD80, CD83, CD1a and HLA DR on the DCs surface in SIBO positive cirrhotic patients were lower than those in SIBO negative [(17.22 ± 2.08) % vs (22.13 ± 2.34) %, (24.65 ± 1.47) % vs ( 34.52 ± 2.84) %, (14.05 ± 1.33) % vs (17.40 ± 3.22) %, (81.47 ± 7.56)% vs (70.15 ± 6.32) %; (P < 0.05)]. Conclusions: The prevalence of SIBO is correlated with severity of cirrhosis. The phenotype and function of peripheral blood DCs in cirrhotic patients with hepatitis B is associated with the presence of SIBO.

Hypoxia-inducible factor-1a contributes to dendritic overgrowth in tuberous sclerosis

Expression of hypoxia-inducible factor 1a (HIF1a) is increased under several pathological conditions such as hyperactive mechanistic target of rapamycin complex 1 (mTORC1) in tuberous sclerosis complex (TSC). Hyperactive mTORC1 and the resulting increased dendritic complexity of neurons are shared molecular and cellular alterations in several neurological disorders associated with cognitive disabilities. Despite some evidence that HIF1a contributes to dendritic overgrowth in vitro, it remains unknown whether increased HIF1a in TSC neurons could contribute to their increased dendritic complexity. To address this use in vivo, we generated TSC neurons by deleting Tsc1 in newborn olfactory bulb (OB) neurons of conditional Tsc1 transgenic mice using neonatal electroporation. In addition to their increased dendritic complexity, Tsc1null neurons have been reported to display increased Hif1a mRNA level and HIF1a transcriptional activity. We found that Tsc1null-dependent dendritic overgrowth was prevented by knocking down HIF1a or expressing a dominant negative HIF1a. In addition, overexpressing HIF1a in wild-type developing neurons resulted in increased dendritic complexity in vivo. These data highlight that an increase in HIF1a levels contributes to abnormal dendritic patterning in developing neurons under normal conditions and hyperactive mTORC1 conditions as in TSC.

Prenatal Deletion of the RNA-Binding Protein HuD Disrupts Postnatal Cortical Circuit Maturation and Behavior

The proper functions of cortical circuits are dependent upon both appropriate neuronal subtype specification and their maturation to receive appropriate signaling. These events establish a balanced circuit that is important for learning, memory, emotion, and complex motor behaviors. Recent research points to mRNA metabolism as a key regulator of this development and maturation process. Hu antigen D (HuD), an RNA-binding protein, has been implicated in the establishment of neuronal identity and neurite outgrowth in vitro. Therefore, we investigated the role of HuD loss of function on neuron specification and dendritogenesis in vivo using a mouse model. We found that loss of HuD early in development results in a defective early dendritic overgrowth phase and pervasive deficits in neuron specification in the lower neocortical layers and defects in dendritogenesis in the CA3 region of the hippocampus. Subsequent behavioral analysis revealed a deficit in performance of a hippocampus-dependent task: the Morris water maze. Further, HuD knock-out (KO) mice exhibited lower levels of anxiety than their wild-type counterparts and were overall less active. Last, we found that HuD KO mice are more susceptible to auditory-induced seizures, often resulting in death. Our findings suggest that HuD is necessary for the establishment of neocortical and hippocampal circuitry and is critical for their function.

Dendritic Arborization and Spine Dynamics Are Abnormal in the Mouse Model ofMECP2Duplication Syndrome

MECP2 duplication syndrome is a childhood neurological disorder characterized by intellectual disability, autism, motor abnormalities, and epilepsy. The disorder is caused by duplications spanning the gene encoding methyl-CpG-binding protein-2 (MeCP2), a protein involved in the modulation of chromatin and gene expression. MeCP2 is thought to play a role in maintaining the structural integrity of neuronal circuits. Loss of MeCP2 function causes Rett syndrome and results in abnormal dendritic spine morphology and decreased pyramidal dendritic arbor complexity and spine density. The consequences of MeCP2 overexpression on dendritic pathophysiology remain unclear. We used in vivo two-photon microscopy to characterize layer 5 pyramidal neuron spine turnover and dendritic arborization as a function of age in transgenic mice expressing the human MECP2 gene at twice the normal levels of MeCP2 (Tg1; Collins et al., 2004). We found that spine density in terminal dendritic branches is initially higher in young Tg1 mice but falls below control levels after postnatal week 12, approximately correlating with the onset of behavioral symptoms. Spontaneous spine turnover rates remain high in older Tg1 animals compared with controls, reflecting the persistence of an immature state. Both spine gain and loss rates are higher, with a net bias in favor of spine elimination. Apical dendritic arbors in both simple- and complex-tufted layer 5 Tg1 pyramidal neurons have more branches of higher order, indicating that MeCP2 overexpression induces dendritic overgrowth. P70S6K was hyperphosphorylated in Tg1 somatosensory cortex, suggesting that elevated mTOR signaling may underlie the observed increase in spine turnover and dendritic growth.

PTEN regulates retinal interneuron morphogenesis and synaptic layer formation

The lipid phosphatase PTEN is a critical negative regulator of extracellular signal-induced PI3K activities, yet the roles of PTEN in the neural retina remain poorly understood. Here, we investigate the function of PTEN during retinal development. Deletion of Pten at the onset of neurogenesis in retinal progenitors results in the reduction of retinal ganglion cells and rod photoreceptors, but increased Müller glial genesis. In addition, PTEN deficiency leads to elevated phosphorylation of Akt, especially in the developing inner plexiform layer, where high levels of PTEN are normally expressed. In Pten mutant retinas, various subtypes of amacrine cells show severe dendritic overgrowth, causing specific expansion of the inner plexiform layer. However, the outer plexiform layer remains relatively undisturbed in the Pten deficient retina. Physiological analysis detects reduced rod function and augmented oscillatory potentials originating from amacrine cells in Pten mutants. Furthermore, deleting Pten or elevating Akt activity in individual amacrine cells is sufficient to disrupt dendritic arborization, indicating that Pten activity is required cell autonomously to control neuronal morphology. Moreover, inhibiting endogenous Akt activity attenuates inner plexiform layer formation in vitro. Together, these findings demonstrate that suppression of PI3K/Akt signaling by PTEN is crucial for proper neuronal differentiation and normal retinal network formation.

PTX-induced hyperexcitability affects dendritic shape and GABAergic synapse density but not synapse distribution during Manduca postembryonic motoneuron development

During the metamorphosis of the holometabolous insect, Manduca sexta, the postembryonic acquisition of adult specific motor behaviors is accompanied by changes in dendritic architecture, membrane currents, and input synapses of identified motoneurons. This study aims to test whether increased activity affects dendritic architecture and sub-dendritic input synapse distribution of the identified flight motoneuron 5 (MN5). Systemic injections of the chloride channel blocker, picrotoxin (PTX), during early pupal stages increase pupal reflex responsiveness, but overall development is not impaired. MN5 input resistance, resting membrane potential, and spiking threshold are not affected. Bath application of PTX to isolated ventral nerve cords evokes spiking in pupal and adult flight motoneurons. Quantitative three-dimensional reconstructions of the dendritic tree of the adult MN5 show that systemic PTX injections into early pupae cause dendritic overgrowth and reduce the density of GABAergic inputs. In contrast, the distribution patterns of GABAergic terminals throughout the dendritic tree remain unaltered. This indicates that increased overall excitability might cause dendritic overgrowth and decreased inhibitory input during postembryonic motoneuron remodeling, whereas sub-dendritic synapse targeting might be controlled by activity-independent signals. Behavioral testing reveals that these neuronal changes do not impede the animal's ability to fly, but impair maximum flight performance.

Bdnf overexpression in hippocampal neurons prevents dendritic atrophy caused by Rett-associated MECP2 mutations

The expression of the methylated DNA-binding protein MeCP2 increases during neuronal development, which suggests that this epigenetic factor is crucial for neuronal terminal differentiation. We evaluated dendritic and axonal development in embryonic day-18 hippocampal neurons in culture by measuring total length and counting branch point numbers at 4 days in vitro, well before synapse formation. Pyramidal neurons transfected with a plasmid encoding a small hairpin RNA (shRNA) to knockdown endogenous Mecp2 had shorter dendrites than control untransfected neurons, without detectable changes in axonal morphology. On the other hand, overexpression of wildtype (wt) human MECP2 increased dendritic branching, in addition to axonal branching and length. Consistent with reduced neuronal growth and complexity in Rett syndrome (RTT) brains, overexpression of human MECP2 carrying missense mutations common in RTT individuals (R106W or T158M) reduced dendritic and axonal length. One of the targets of MeCP2 transcriptional control is the Bdnf gene. Indeed, endogenous Mecp2 knockdown increased the intracellular levels of BDNF protein compared to untransfected neurons, suggesting that MeCP2 represses Bdnf transcription. Surprisingly, overexpression of wt MECP2 also increased BDNF levels, while overexpression of RTT-associated MECP2 mutants failed to affect BDNF levels. The extracellular BDNF scavenger TrkB-Fc prevented dendritic overgrowth in wt MECP2-overexpressing neurons, while overexpression of the Bdnf gene reverted the dendritic atrophy caused by Mecp2-knockdown. However, this effect was only partial, since Bdnf increased dendritic length only to control levels in mutant MECP2-overexpressing neurons, but not as much as in Bdnf-transfected cells. Our results demonstrate that MeCP2 plays varied roles in dendritic and axonal development during neuronal terminal differentiation, and that some of these effects are mediated by autocrine actions of BDNF.

Neuronal Homeostasis: Does Form Follow Function or Vice Versa?

Nerve cells adjust their electrical excitability and the overall strength of synaptic connections to maintain their functional identity. A recent study suggests that they may also regulate dendritic branch patterns to compensate for the variability of synaptic contacts and help ensure appropriate connectivity in the brain.

Dendrite Elongation and Dendritic Branching Are Affected Separately by Different Forms of Intrinsic Motoneuron Excitability

Dendrites are the fundamental determinant of neuronal wiring. Consequently dendritic defects are associated with numerous neurological diseases and mental retardation. Neuronal activity can have profound effects on dendritic structure, but the mechanisms controlling distinct aspects of dendritic architecture are not fully understood. We use the Drosophila genetic model system to test the effects of altered intrinsic excitability on postembryonic dendritic architecture development. Targeted dominant negative knock-downs of potassium channel subunits allow for selectively increasing the intrinsic excitability of a selected subset of motoneurons, whereas targeted expression of a genetically modified noninactivating potassium channel decrease intrinsic excitability in vivo. Both manipulations cause significant dendritic overgrowth, but by different mechanisms. Increased excitability causes increased dendritic branch formation, whereas decreased excitability causes increased dendritic branch elongation. Therefore dendritic branching and branch elongation are controlled by separate mechanisms that can be addressed selectively in vivo by different manipulations of neuronal intrinsic excitability.

Dendritic overgrowth and alterations in laminar phenotypes of neocortical neurons in the newborn with semilobar holoprosencephaly

We analyzed neuronal phenotypes and dendritic growth in the newborn with semilobar holoprosencephaly and 18p deletion. We found that the holoprosencephalic neocortex retained its basic six-layered lamination but displayed a number of intralaminar and modular architectonic alterations and contained a mixture of normal and aberrant neuronal phenotypes. The most conspicious finding was the presence of the pronounced increase in soma size and total basal dendritic length of holoprosencephalic layer III pyramidal neurons in comparison to age-matched control brains. The dramatic (5-fold) dendritic overgrowth observed in associative cortico-cortical pyramidal neurons is probably related to the pronounced diminution of the cortical afferent input.

Use-dependent growth of pyramidal neurons after neocortical damage

Unilateral damage to the forelimb representation area of the sensorimotor cortex in adult rats increases dendritic arborization of layer V pyramidal neurons of the contralateral homotopic cortex. Arbor size was maximum at approximately 18 d postlesion, following which there was a partial elimination, or pruning, of dendritic processes. These neural changes were closely associated with behavioral events. The overgrowth of dendrites was related in time to disuse of the contralateral (to the lesion) forelimb and over-reliance on the ipsilateral forelimb for postural and exploratory movements. The pruning of dendrites was related to a return to more symmetrical use of the forelimbs. To investigate the possibility that lesion-induced asymmetries in motor behavior contributed to dendritic arborization changes, movements of the forelimb ipsilateral to the lesion were restricted during the period of dendritic overgrowth through the use of one-holed vests. This interfered with the increase in dendritic arborization. In contrast, animals that were allowed to use both forelimbs, or only the forelimb ipsilateral to the lesion, showed the expected increases. When sham-operated rats were forced to use only one forelimb, no significant increases in arborization were found. Therefore, neither a lesion nor asymmetrical limb use alone could account for the dendritic overgrowth--it depended on a lesion-behavior interaction. Furthermore, greater sensorimotor impairments were found when the dendritic growth was blocked, suggesting that the neural growth and/or associated limb-use behavior were related to functional recovery from the cortical damage. Finally, in a second experiment, immobilization of the impaired limb during the pruning period did not prevent the elimination of processes. Thus, the pruning of neural processes was not related simply to the recovery of more symmetrical forelimb use. There may be a period early after brain damage during which marked neural structural changes can occur in the presence of adequate behavioral demand.