The confinement of neuronal activity to specific subcellular regions is a mechanism for expanding the computational properties of neurons. Although the circuit organization underlying compartmentalized activity has been studied in several systems, its cellular basis is still unknown. Here we characterize compartmentalized activity in Caenorhabditis elegans RIA interneurons, which have multiple reciprocal connections to head motor neurons and receive input from sensory pathways. We show that RIA spatially encodes head movement on a subcellular scale through axonal compartmentalization. This subcellular axonal activity is dependent on acetylcholine release from head motor neurons and is simultaneously present and additive with glutamate-dependent globally synchronized activity evoked by sensory inputs. Postsynaptically, the muscarinic acetylcholine receptor GAR-3 acts in RIA to compartmentalize axonal activity through the mobilization of intracellular calcium stores. The compartmentalized activity functions independently of the synchronized activity to modulate locomotory behaviour.
Many animals use their olfactory systems to learn to avoid dangers, but how neural circuits encode naive and learned olfactory preferences, and switch between those preferences, is poorly understood. Here, we map an olfactory network, from sensory input to motor output, which regulates the learned olfactory aversion of Caenorhabditis elegans for the smell of pathogenic bacteria. Naive animals prefer smells of pathogens but animals trained with pathogens lose this attraction. We find that two different neural circuits subserve these preferences, with one required for the naive preference and the other specifically for the learned preference. Calcium imaging and behavioral analysis reveal that the naive preference reflects the direct transduction of the activity of olfactory sensory neurons into motor response, whereas the learned preference involves modulations to signal transduction to downstream neurons to alter motor response. Thus, two different neural circuits regulate a behavioral switch between naive and learned olfactory preferences.
Axonal growth cones use intermediate targets to navigate in the developing nervous system. Encountering these sites is correlated with growth cone pausing. PHR (Phr1, Esrom, Highwire, RPM-1) is a large neuronal ubiquitin ligase that interacts with multiple signaling pathways. Mouse and zebrafish phr mutants have highly penetrant axon pathfinding defects at intermediate targets. Mouse phr mutants contain excessive microtubules in the growth cone, which has been attributed to upregulation of DLK/p38 signaling. Here, we ask whether this pathway and microtubule misregulation are indeed linked to guidance errors in the vertebrate brain, using the zebrafish. By live imaging, we show that loops form when microtubules retract without depolymerizing. JNK, but not p38, phosphorylation is increased in mutant growth cones. However microtubule looping cannot be suppressed by inhibiting JNK. The phr microtubule defect can be phenocopied by taxol, while microtubule destabilization in vitro using nocodazole prevents loop formation. Acute disruption in vivo with nocodazole suppresses the intermediate target guidance defect. Given that microtubule looping is associated with growth cone pausing, we propose that microtubule disassembly, mediated by PHR, is essential for exiting the paused state at intermediate targets.
Growth cones are guided to their final destination by intermediate targets. Here, we identify intermediate targets and signaling components acting on zebrafish habenula commissural axons. Live imaging establishes that axons pause at the medial habenula before and after crossing the roof plate. esrom mutants axons fail to advance beyond the ipsilateral medial habenula. Tsc2 function is reduced in mutant axons, indicating cell autonomous defects in signaling. Consistent with signaling properties changing outside the roof plate, EphB is surface localized on axon segments within a zone demarcated by the medial habenula. wnt4a is expressed in the medial habenula and morpholino knockdown causes loss of the commissure. Electroporation of truncated Ryk causes axons to reenter the midline after reaching the contralateral habenula. These data identify Esrom as a mediator of growth cone navigation at an intermediate target and underscore the importance of midline boundaries as signaling centers for commissure formation.
The habenular complex is a paired structure found in the diencephalon of all vertebrates, linking the forebrain and midbrain. Habenulae are asymmetrical and may contribute to lateralized behavior. Recent studies in zebrafish have characterized molecular pathways that give rise to the habenular asymmetry and the distinct projections of the left and right habenula to the midbrain. However, it is unclear whether there are asymmetries in habenula afferents from the forebrain. By lipophilic dye tracing, we find that axons innervating the habenula derive primarily from a region in the lateral diencephalon containing migrated neurons of the eminentia thalami (EmT). EmT neurons terminate in neuropils in both ipsilateral and contralateral habenula. These axons, together with axons from migrated neurons of the posterior tuberculum and pallial neurons, cross the midline via the habenular commissure. Subsets of pallial neurons terminate only in the medial right habenula, regardless of which side of the brain they originate from. These include an unusual type of forebrain projection: axons that cross the midline twice, at both the anterior and habenular commissures. Our data establish that there is asymmetric innervation of the habenula from the telencephalon, suggesting a mechanism by which habenula asymmetry might contribute to lateralized behavior.
Electroporation is a technique for the introduction of nucleic acids and other macromolecules into cells. In chick embryos it has been a particularly powerful technique for the spatial and temporal control of gene expression in developmental studies. Electroporation methods have also been reported for Xenopus, zebrafish, and mouse.
Visual system development is dependent on correct interpretation of cues that direct growth cone migration and axon branching. Mutations in the zebrafish esrom gene disrupt bundling and targeting of retinal axons, and also cause ectopic arborization. By positional cloning, we establish that esrom encodes a very large protein orthologous to PAM (protein associated with Myc)/Highwire/RPM-1. Unlike motoneurons in Drosophila highwire mutants, retinal axons in esrom mutants do not arborize excessively, indicating that Esrom has different functions in the vertebrate visual system. We show here that Esrom has E3 ligase activity and modulates the amount of phosphorylated Tuberin, a tumor suppressor, in growth cones. These data identify a mediator of signal transduction in retinal growth cones, which is required for topographic map formation.