Axonal branch patterning and neuronal shape diversity: roles in developmental circuit assembly: Axonal branch patterning and neuronal shape diversity in developmental circuit assembly

Recent progress in human genetics and single cell sequencing rapidly expands the list of molecular factors that offer important new contributions to our understanding of brain wiring. Yet many new molecular factors are being discovered that have never been studied in the context of neuronal circuit development. This is clearly asking for increased efforts to better understand the developmental mechanisms of circuit assembly [1]. Moreover, recent studies characterizing the developmental causes of some psychiatric diseases show impressive progress in reaching cellular resolution in their analysis. They provide concrete support emphasizing the importance of axonal branching and synapse formation as a hotspot for potential defects. Inspired by these new studies we will discuss progress but also challenges in understanding how neurite branching and neuronal shape diversity itself impacts on specificity of neuronal circuit assembly. We discuss the idea that neuronal shape acquisition itself is a key specificity factor in neuronal circuit assembly.     

Axis formation and patterning in zebrafish.

A large collection of mutations affecting zebrafish embryogenesis was described in 1996. The cloning of the affected genes has now provided novel insights into the role and regulation of signaling by BMP, Nodal, Wnt, FGF, Hedgehog, Delta, Slit, retinoic acid and lipids. Detailed analyses have revealed a complex genetic network that patterns the early embryo.     

The formation and patterning of leaves: recent advances

Leaves, the plants major photosynthetic organs, form through the activity of groups of pluripotent cells, termed shoot apical meristems (SAMs), located at the growing tips of plants. Leaves develop with a dorso–ventral asymmetry, with the adaxial surface adjacent to the meristem and the abaxial surface developing at a distance from it. Molecular genetic studies have shown that the correct specification of adaxial/abaxial polarity requires communication between the incipient leaf and the meristem, and that the juxtaposition of adaxial/abaxial fates is necessary for lamina outgrowth (Waites and Hudson 1995; McConnell et al. 2001). Over the last few years, a number of factors that control cell fate specification in the apex have been identified. This review will focus on recent advances on distinct but overlapping aspects of leaf development, namely, the transition from meristem to leaf fate and the specification of abaxial/adaxial polarity and its possible role in leaf growth.     

Neuroblast formation and patterning during early brain development in Drosophila

The Drosophila embryo provides a useful model system to study the mechanisms that lead to pattern and cell diversity in the central nervous system (CNS). The Drosophila CNS, which encompasses the brain and the ventral nerve cord, develops from a bilaterally symmetrical neuroectoderm, which gives rise to neural stem cells, called neuroblasts. The structure of the embryonic ventral nerve cord is relatively simple, consisting of a sequence of repeated segmental units (neuromeres), and the mechanisms controlling the formation and specification of the neuroblasts that form these neuromeres are quite well understood. Owing to the much higher complexity and hidden segmental organization of the brain, our understanding of its development is still rudimentary. Recent investigations on the expression and function of proneural genes, segmentation genes, dorsoventral-patterning genes and a number of other genes have provided new insight into the principles of neuroblast formation and patterning during embryonic development of the fly brain. Comparisons with the same processes in the trunk help us to understand what makes the brain different from the ventral nerve cord. Several parallels in early brain patterning between the fly and the vertebrate systems have become evident.     

Theoretical models for branch formation in plants

Various branch architectures are observed in living organisms including plants. Branch formation has traditionally been an area of interest in the field of developmental biology, and theoretical approaches are now commonly used to understand the complex mechanisms involved. In this review article, we provide an overview of theoretical approaches including mathematical models and computer simulations for studying plant branch formation. These approaches cover a wide range of topics. In particular, we focus on the importance of positional information in branch formation, which has been especially revealed by theoretical research in plants including computations of developmental processes.

     

Disruption of early proximodistal patterning and AVE formation in Apc mutants

In the postimplantation mouse embryo, axial patterning begins with the restriction of expression of a set of genes to the distal visceral endoderm (DVE). This proximodistal (PD) axis is subsequently transformed into an anteroposterior axis as the VE migrates anteriorly to form the anterior visceral endoderm (AVE). Both Nodal and Wnt signaling pathways are involved in these events. We show here that loss of function in the adenomatous polyposis coli gene (Apc) leads to constitutive beta-catenin activity that induces a proximalization of the epiblast with the activation of a subset of posterior mesendodermal genes, and loss of ability to induce the DVE. The loss of some DVE genes such as Hex and goosecoid is rescued in chimeras where only the epiblast was wild type; however, these DVE markers were no longer restricted distally but covered the entire epiblast. Thus, the Apc gene is needed in both embryonic and extraembryonic lineages for normal PD patterning around implantation, suggesting that early restricted activation of the Wnt pathway may be important for initiating axial asymmetries. In addition, we found that nuclear beta-catenin and other molecular markers are asymmetrically expressed by 4.5 days.     

Phytochrome-mediated branch formation in protonemata of the mossCeratodon purpureus

We have analyzed light induction of side-branch formation and chloroplast re-arrangement in protonemata of the mossCeratodon purpureus. After 12 hr of dark adaptation, the rate of branch formation was as low as 5%. A red light treatment induced formation of side branches up to 75% of the dark-adapted protonema. The frequency of light induced branch formation differed between cells of different ages, the highest frequency being found in the 5th cell, the most distal cell studied from the apex. We examined the effect of polarized light given parallel to the direction of filament growth. The position of branching within the cell depended on the vibration plane of polarized red light. Branch formation was highest when the electric vector of polarized light vibrates parallel to the cell surface and is fluence rate dependent. The positional effect of polarized red light could be nullified to some extent by simultaneous irradiation with polarized far-red light. An aphototropic mutant,ptr116, shows characteristics of deficiency in biosynthesis of the phytochrome chromophore and exhibits no red-light induced branch formation. Biliverdin, a precursor of the phytochrome chromophore, rescued the red-light induced branching when added to the medium, supporting the conclusion that phytochrome acts as photoreceptor for red light induced branch formation. The light effect on chloroplast re-arrangement was also analyzed in this study. We found that polarized blue light induced chloroplast re-arrangement in wild-type cells, whereas polarized red light was inactive. This result suggests that chloroplast re-arrangement is only controlled by a blue light photoreceptor, not by phytochrome inCeratodon.     

Extra spike formation in sensory neurons and the disruption of afferent spike patterning

The peculiar pseudounipolar geometry of primary sensory neurons can lead to ectopic generation of "extra spikes" in the region of the dorsal root ganglion potentially disrupting the fidelity of afferent signaling. We have used an explicit model of myelinated vertebrate sensory neurons to investigate the location and mechanism of extra spike formation, and its consequences for distortion of afferent impulse patterning. Extra spikes originate in the initial segment axon under conditions in which the soma spike becomes delayed and broadened. The broadened soma spike then re-excites membrane it has just passed over, initiating an extra spike which propagates outwards into the main conducting axon. Extra spike formation depends on cell geometry, electrical excitability, and the recent history of impulse activity. Extra spikes add to the impulse barrage traveling toward the spinal cord, but they also travel antidromically in the peripheral nerve colliding with and occluding normal orthodromic spikes. As a result there is no net increase in afferent spike number. However, extra spikes render firing more staccato by increasing the number of short and long interspike intervals in the train at the expense of intermediate intervals. There may also be more complex changes in the pattern of afferent spike trains, and hence in afferent signaling.     

Regeneration in Medusa buds and Medusae of Hydrozoa

Isolated medusa buds of Podocoryne carnea (Anthomedusa) caneither develop autonomously to functional medusae or transformto stolons and polyps. Up to stage 8 the percentage of transformaiioncan be idised 10 100% by repeated traumatization of the isolatedbuds. From stage 9 onwards, even traumatized buds always regeneratemedusae and never transform to polyps. At stage 8 the bud isdifferentiated, lacking only the swelling of the outer mesogleato become a normal medusa. A correlation between the swellingof this mesoglea and the decrease of the percentage of transformationto zero can be observed between stages 8 and 9. The presenceof a swollen mesoglea is not the only factor in stabilizingmedusa structures, since, despite removal of the mesoglea bythe dissociation method, aggregates of dissociated buds of stages7 and 8 always formed medusa structures and never transformedto stolons or polyps. Regeneration of the manubrium in adult Hydromedusae shows agraded distribution, being highest in the center of the umbrellaand decreasing rapidly towards the circular canal. The extentof manubrium regeneration is not influenced by the amount ofwound material or the components of the gastrovascular system.     

Neural crest and placode roles in formation and patterning of cranial sensory ganglia in lamprey

Vertebrates possess paired cranial sensory ganglia derived from two embryonic cell populations, neural crest and placodes. Cranial sensory ganglia arose prior to the divergence of jawed and jawless vertebrates, but the developmental mechanisms that facilitated their evolution are unknown. Using gene expression and cell lineage tracing experiments in embryos of the sea lamprey, Petromyzon marinus, we find that in the cranial ganglia we targeted, development consists of placode‐derived neuron clusters in the core of ganglia, with neural crest cells mostly surrounding these neuronal clusters. To dissect functional roles of neural crest and placode cell associations in these developing cranial ganglia, we used CRISPR/Cas9 gene editing experiments to target genes critical for the development of each population. Genetic ablation of SoxE2 and FoxD‐A in neural crest cells resulted in differentiated cranial sensory neurons with abnormal morphologies, whereas deletion of DlxB in cranial placodes resulted in near‐total loss of cranial sensory neurons. Taken together, our cell‐lineage, gene expression, and gene editing results suggest that cranial neural crest cells may not be required for cranial ganglia specification but are essential for shaping the morphology of these sensory structures. We propose that the association of neural crest and placodes in the head of early vertebrates was a key step in the organization of neurons and glia into paired sensory ganglia.     

Bottle cell formation in relation to mesodermal patterning in the Xenopus embryo

The appearance of bottle cells at the dorsal vegetal/marginal boundary of Xenopus embryos marks the onset of blastopore formation. The conditions leading to this epithelial activity were investigated by inducing bottle cells ectopically in the animal region with VegT or different members of the transforming growth factor (TGF)-beta family. Morphological studies on the ectopic bottle cells indicate their close similarity to the endogenous bottle cells at the dorsal blastopore lip. The subepithelial cells of the induced animal region express mesodermal genes in a pattern reminiscent to that observed on the dorsal lip. Relating this expression pattern to the position of the ectopic bottle cells leads to the conclusion that bottle cells form in regions of high TGF-beta signalling. The specific inhibitory effects of cerberus on ectopically induced bottle cells revealed that nodal related growth factors are the intrinsic signals that elicit bottle cell formation in the normal embryo. In addition, fibroblast growth factor signalling is an essential precondition for this epithelial response as it is for mesoderm formation. We conclude that bottle cell formation in the epithelial layer of the gastrula is closely linked to mesodermal patterning in the subepithelial tissues.     

Principles of branch dynamics governing shape characteristics of cerebellar Purkinje cell dendrites

Neurons develop dendritic arbors in cell type-specific patterns. Using growing Purkinje cells in culture as a model, we performed a long-term time-lapse observation of dendrite branch dynamics to understand the rules that govern the characteristic space-filling dendrites. We found that dendrite architecture was sculpted by a combination of reproducible dynamic processes, including constant tip elongation, stochastic terminal branching, and retraction triggered by contacts between growing dendrites. Inhibition of protein kinase C/protein kinase D signaling prevented branch retraction and significantly altered the characteristic morphology of long proximal segments. A computer simulation of dendrite branch dynamics using simple parameters from experimental measurements reproduced the time-dependent changes in the dendrite configuration in live Purkinje cells. Furthermore, perturbation analysis to parameters in silico validated the important contribution of dendritic retraction in the formation of the characteristic morphology. We present an approach using live imaging and computer simulations to clarify the fundamental mechanisms of dendrite patterning in the developing brain.     

Cytoskeletal pattern changes during branch formation in a centrifugedAdiantum protonema

A protonemal branch was induced on a side wall of a fern filamentous protonema by cell centrifugation and subsequent polarized-red light irradiation as described in a previous paper (Wada 1995, J. Plant Res. 108: 501–509). Changes in microtubule (MT) and microfilament (MF) patters during the branch development were observed under fluorescence microscopy. A ring-like band of cortical MTs (MT-ring) and MFs similar to a preprophase band or a subapical ring structure (Murataet al. 1987) appeared transiently at the future branching site before cell swelling, the first visible step of branch formation. At this stage, the nucleus was located far from the branching site and the MT-ring appeared to be connected to the nucleus by endoplasmic MFs as well as with endoplasmic MTs. The MT-ring disappeared when cell wall swelling occurred. When the cell wall swelling began, a fan-like pattern of cortical MTs emanating from the new growing tip was established and the MTs reached the opposite flank of the protonema. When a new branch started to elongate and the nucleus moved into the branch, a faint subapical ring of MTs appeared at the subapical part of the new branch. Strands of MTs and MFs emanating from the nuclear front end reached a part of the subapical ring.