Individual Axon Arbors Research Articles

Determinants of synaptic strength vary across an axon arbor

We used synaptophysin-pHluorin expressed in hippocampal neurons to address how functional properties of terminals, namely, evoked release, total vesicle pool size, and release fraction, vary spatially across individual axon arbors. Consistent with previous reports, over short arbor distances (≈ 100 μm), evoked release was spatially heterogeneous when terminals contacted different postsynaptic dendrites or neurons. Regardless of the postsynaptic configuration, the evoked release and total vesicle pool size spatially covaried, suggesting that the fraction of synaptic vesicles available for release (release fraction) was similar over short distances. Evoked release and total vesicle pool size were highly correlated with the amount of NMDA receptors and PSD-95 in postsynaptic specialization. However, when individual axons were followed over longer distances (several hundred micrometers), a significant increase in evoked release was observed distally that was associated with an increased release fraction in distal terminals. The increase in distal release fraction can be accounted for by changes in individual vesicle release probability as well as readily releasable pool size. Our results suggest that for a single axon arbor, presynaptic strength indicated by evoked release over short distances is correlated with heterogeneity in total vesicle pool size, whereas over longer distances presynaptic strength is correlated with the spatial modulation of release fraction. Thus the mechanisms that determine synaptic strength differ depending on spatial scale.

Neocortical Axon Arbors Trade-off Material and Conduction Delay Conservation

The brain contains a complex network of axons rapidly communicating information between billions of synaptically connected neurons. The morphology of individual axons, therefore, defines the course of information flow within the brain. More than a century ago, Ramón y Cajal proposed that conservation laws to save material (wire) length and limit conduction delay regulate the design of individual axon arbors in cerebral cortex. Yet the spatial and temporal communication costs of single neocortical axons remain undefined. Here, using reconstructions of in vivo labelled excitatory spiny cell and inhibitory basket cell intracortical axons combined with a variety of graph optimization algorithms, we empirically investigated Cajal's conservation laws in cerebral cortex for whole three-dimensional (3D) axon arbors, to our knowledge the first study of its kind. We found intracortical axons were significantly longer than optimal. The temporal cost of cortical axons was also suboptimal though far superior to wire-minimized arbors. We discovered that cortical axon branching appears to promote a low temporal dispersion of axonal latencies and a tight relationship between cortical distance and axonal latency. In addition, inhibitory basket cell axonal latencies may occur within a much narrower temporal window than excitatory spiny cell axons, which may help boost signal detection. Thus, to optimize neuronal network communication we find that a modest excess of axonal wire is traded-off to enhance arbor temporal economy and precision. Our results offer insight into the principles of brain organization and communication in and development of grey matter, where temporal precision is a crucial prerequisite for coincidence detection, synchronization and rapid network oscillations.

Coordinated Motor Neuron Axon Growth and Neuromuscular Synaptogenesis Are Promoted by CPG15 In Vivo

We have used in vivo time-lapse two-photon imaging of single motor neuron axons labeled with GFP combined with labeling of presynaptic vesicle clusters and postsynaptic acetylcholine receptors in Xenopus laevis tadpoles to determine the dynamic rearrangement of individual axon branches and synaptogenesis during motor axon arbor development. Control GFP-labeled axons are highly dynamic during the period when axon arbors are elaborating. Axon branches emerge from sites of synaptic vesicle clusters. These data indicate that motor neuron axon elaboration and synaptogenesis are concurrent and iterative. We tested the role of Candidate Plasticity Gene 15 (CPG15, also known as Neuritin), an activity-regulated gene that is expressed in the developing motor neurons in this process. CPG15 expression enhances the development of motor neuron axon arbors by promoting neuromuscular synaptogenesis and by increasing the addition of new axon branches.

Development of Individual Axon Arbors in a Thalamocortical Circuit Necessary for Song Learning in Zebra Finches

Individual axon arbors within developing neural circuits are remodeled during restricted sensitive periods, leading to the emergence of precise patterns of connectivity and specialized adaptive behaviors. In male zebra finches, the circuit connecting the medial dorsolateral nucleus of the thalamus (DLM) and its cortical target, the lateral magnocellular nucleus of the anterior neostriatum (lMAN), is crucial for the acquisition of a normal vocal pattern during the sensitive period for song learning. The shell subregion of lMAN as well as the entire terminal field of DLM axons within lMAN undergo a striking increase in overall volume during early stages of vocal learning followed by an equally substantial decrease by adulthood, by which time birds have acquired stable song patterns. Because the total number of DLM neurons remains stable throughout this period, the dramatic changes within the overall DLM-->lMAN circuit are presumably attributable to dynamic rearrangements at the level of individual DLM axon arbors over the course of vocal learning. To study such rearrangements directly, we reconstructed individual DLM axon arbors in three dimensions at different stages during vocal learning. Unlike axon arbors in other model systems, in which the number of branches increases during development, DLM arbors are unusual in that they have the greatest number of branches at the onset of vocal learning and undergo large-scale retraction during the sensitive period for song learning. Decreases in the degree of overlap between DLM arbors apparently contribute to the increased overall volume of the DLM-->lMAN circuit during vocal learning. These developmental changes in DLM axon arbors occur at the height of the sensitive period for vocal learning, and hence may represent either a morphological correlate of song learning or a necessary prerequisite for acquisition of song.

Individual axon morphology and thalamocortical topography in developing rat somatosensory cortex.

The morphology of individual thalamocortical axons in developing rat primary somatosensory cortex was studied using lipophilic tracers. Anterograde labeling with lipophilic dyes demonstrated a topographical organization of thalamocortical projections exiting the thalamus as early as embryonic day (E) 16; retrograde labeling studies demonstrated topography of these projections as they reached the cortex as early as E18. At E17, axons course tangentially within the intermediate zone and turn or branch near the deepest layer of cortex (layer VIb), suggesting the presence of guidance cues in this region. Axons appear to grow and branch progressively within layers VIb and VIa during the following days; axons in the intermediate zone may give rise to radially directed branches. Individual axons appear to grow steadily and progressively into the cortex, with the leading front of axons at the transition zone between the cortical plate (CP) and the differentiating cortical layers. At birth (P0), thalamocortical axons extend radially through layers VIa and V and emit branches within these layers; some axons reach the CP. By P1, layer IV has begun to differentiate and axons begin to form a few simple branches in the vicinity of the layer IV cells. Over the ensuing week, axons generate more branches within layer IV, but the tangential extent of individual axon arbors does not exceed the width of a barrel. By P7, individual axons overlap within barrel clusters, and individual axons span the width of a cluster. These observations indicate that thalamic afferents develop by progressive growth of arbors that remain spatially restricted, rather than by overbranching and retracting arbors.

Projection of individual axons from the pretectum to the dorsal lateral geniculate complex in the cat.

The dorsal lateral geniculate nucleus of the thalamus transmits visual signals from the retina to the cortex. Within the lateral geniculate nucleus, the ascending visual signals are modified by the actions of a number of afferent pathways. One such projection originates in the pretectum and appears to be active in association with oculomotor activity. Much remains unknown about the pretectal-geniculate projection. Our purpose was to examine for the first time individual axon arbors from the pretectum that project to the lateral geniculate nucleus, describing their topography and nuclear and laminar targets. We made injections of the anterograde tracer Phaseolus vulgaris leucoagglutinin into the cat pretectum, targeting the nucleus of the optic tract. Serial 40 microns coronal sections were processed by using immunohistochemistry to reveal labeled axons that were then serially reconstructed using light microscopy. Pretectal-geniculate axons appeared morphologically heterogeneous in terms of swelling size, branching patterns, and laminar target. Most axons innervated the geniculate A laminae. A separate, smaller population innervated the C laminae. All axons exhibited substantially greater spread medial-laterally than rostral-caudally in the lateral geniculate nucleus, displaying a topographical organization for visual field elevation, but not azimuth. Many pretectal axons that projected to the LGN also innervated adjacent structures, including the medial interlaminar nucleus, the perigeniculate nucleus, and/or the pulvinar. These results indicate that the projection from the pretectum to the dorsal lateral geniculate nucleus is heterogeneous, is semitopographical, and may coordinate neural activity in the lateral geniculate nucleus and in neighboring visual thalamic structures in association with oculomotor events.

Regulation of retinal ganglion cell axon arbor size by target availability: mechanisms of compression and expansion of the retinotectal projection.

The ability of pre- and postsynaptic populations to achieve the proper convergence ratios during development is especially critical in topographically mapped systems such as the retinotectal system. The ratio of retinal ganglion cells to their target cells in the optic tectum can be altered experimentally either by early partial tectal ablation, which results in an orderly compression of near-normal numbers of retinal projections into a smaller tectal area, or by early monocular enucleation, which results in the expansion of a reduced number of axons in a near-normal tectal volume. Our previous studies showed that changes in cell death and synaptic density consequent to these manipulations can account for only a minor component of this compensation for the population mismatch. In this study, we examine other mechanisms of population matching in the hamster retinotectal system. We used an in vitro horseradish peroxidase labeling method to trace individual retinal ganglion cell axons in superior colliculi partially ablated on the day of birth, as well as in colliculi contralateral to a monocular enucleation. We found that individual axon arbors within the partially lesioned tectum occupy a smaller area, with fewer branches and fewer terminal boutons, but preserve a normal bouton density. In contrast, ipsilaterally projecting axon arbors in monocularly enucleated animals occupy a greater area than in the normal condition, with a much larger arbor length and greater number of boutons and branches compared with normal ipsilaterally projecting cells. Alteration of axonal arborization of retinal ganglion cells is the main factor responsible for matching the retinal and tectal cell populations within the tectum. This process conserves normal electrophysiological function over a wide range of convergence ratios and may occur through strict selectivity of tectal cells for their normal number of inputs.

Bistratified distribution of terminal arbors of individual axons projecting from area V1 to middle temporal area (MT) in the macaque monkey

In the present study, the anterograde tracer Phaseolus vulgaris-leucoagglutinin was injected into area V1 in order to demonstrate the detailed morphology of individual axons terminating in prestriate area MT. On the basis of 24 axon reconstructions, several representative (but not necessarily comprehensive) characteristics have been identified: (1) Most axons arborize in a patchy manner over a widespread territory, frequently greater than 1.0 mm and often up to 1.5 x 1.8 mm (dimensions uncorrected for shrinkage). (2) Terminal arbors are distributed to layers 3, 4, and 6. Those in layer 6 need not be in register with those in the upper layers. (3) Number and size of terminal arbors are variable. One axon may have 1-4 arbors in the middle layers; typically at least one of these will have a diameter of 200-250 microns, while the others may be less developed. There are from 1-3 arbors in layer 6, usually 50 microns (but sometimes up to 100 microns) in diameter. (4) Terminal boutons are of mixed morphology, but usually beaded and large (up to 3.0 microns). (5) In the white matter, many axons travel in the external sagittal stratum but some are part of the U-fiber system. Axons commonly branch, sometimes at depths up to 0.75-1.0 mm, below the gray matter of MT. In summary, these axons are not stereotyped, but rather vary in the number and size of their terminal arbors, as well as in their branching and overall geometry. Connections from area V1 to MT have been associated with the magnocellular-dominated "processing channel." As widespread arborizations and bistratified terminations are common to both striate axons in MT and to geniculocortical axons in layer 4C alpha of primary visual cortex, these features might be correlated with magnocellular-specific processing requirements.

Abnormal axonal growth in the dorsal lateral geniculate nucleus of the cat.

Retino-geniculate axons in the cat were induced to grow abnormally by cutting one optic nerve in kittens. Surviving optic tract axons that had grown into the denervated regions were then filled in the adults with horseradish peroxidase to reveal the terminal arbors of individual axons. Two types of abnormal axonal growth are described--translaminar growth and monocular segment growth. Translaminar growth is the most common and occurs between laminae in the binocular part to the nucleus. Axons giving rise to translaminar growth do not branch as they pass through the denervated regions of the nucleus. Instead, the abnormal branches originate from portions of the terminal arbor within the normal target lamina. These axons look like normal retino-geniculate axons in terms of their branching patterns, cytological features, and patterns of synaptic contacts except that parts of their terminal arbors have expanded to innervate inappropriate laminae. The distribution of translaminar branches overlaps the distribution of a restricted group of surviving large neurons that have not undergone denervation atrophy. Monocular segment growth invades the lateral pole of the nucleus directly from the optic tract. These branches arise from axons passing through or near the denervated region and appear to represent the formation of new terminal arbors. The synaptic swellings arising from these branches have cytological features like the synaptic swellings arising from translaminar branches and they form similar patterns of synaptic contacts. However, monocular segment branches degenerate more rapidly when damaged and they are not associated with surviving large neurons.