Dynamics of axonal regeneration in adult and aging zebrafish reveal the promoting effect of a first lesion

Axonal regeneration is a major issue in the maintenance of adult nervous systems, both after nerve injuries and in neurodegenerative diseases. However, studying this process in vivo is difficult or even impossible in most vertebrates. Here we show that the posterior lateral line (PLL) of zebrafish is a suitable system to study axonal regeneration in vivo because of both the superficial location and reproducible spatial arrangement of neurons and targets, and the possibility of following reinnervation in live fish on a daily basis. Axonal regeneration after nerve cut has been demonstrated in this system during the first few days of life, leading to complete regeneration within 24 h. However, the potential for PLL nerve regeneration has not been tested yet beyond the early larval stage. We explore the regeneration potential and dynamics of the PLL nerve in adult zebrafish and report that regeneration occurs throughout adulthood. We observed that irregularities in the original branching pattern are faithfully reproduced after regeneration, suggesting that regenerating axons follow the path laid down by the original nerve branches. We quantified the extent of target reinnervation after a nerve cut and found that the latency before the nerve regenerates increases with age. This latency is reduced after a second nerve cut at all ages, suggesting that a regeneration-promoting factor induced by the first cut facilitates regeneration on a second cut. We provide evidence that this factor remains present at the site of the first lesion for several days and is intrinsic to the neurons.The potential of adult neurons to regenerate their axons and to reinnervate target organs after injury is not as well understood as early axonogenesis. Whether and how this capability is modified on aging is an even less explored area. In all vertebrates studied so far, neurons of the peripheral nervous system retain the ability to reextend peripheral axons and reestablish functional connections. This ability is thought to involve intrinsic mechanisms of repair as well as extrinsic signals from the local environment, mostly coming from Schwann cells and macrophages (1).The posterior lateral line (PLL) of fish is a convenient yet unexplored sensory system to address the issue of axonal regeneration throughout adulthood. The PLL comprises a set of superficial mechanosensory organs called neuromasts, which are distributed over the body and tail. Neuromasts are composed of a core of mechanosensory hair cells providing information about the local water flow, surrounded by accessory cells. The afferent neurons innervating neuromasts are clustered in a ganglion posterior to the otic vesicle, and their peripheral axons extend toward the tail, right under the skin. This sensory system is involved in a large repertoire of behaviors (2).The juvenile PLL of zebrafish comprises four lines of neuromasts that extend at different dorsoventral levels, totaling about 50 organs (3). This pattern is established around 1 mo postfertilization (mpf) and remains essentially unchanged throughout adulthood, except that each juvenile neuromast gives rise to a number of “accessory” neuromasts through a budding process (4, 5) that depends on innervation (6). Bud-neuromasts remain closely associated and form dorsoventrally arranged linear clusters, or “stitches” (7). Here we concentrate on the most extensive of the four lines, the ventral one. This line lies originally along the horizontal myoseptum of the embryo and comprises only five neuromasts. More neuromasts develop during larval life, such that the line eventually consists of one neuromast on every intersomitic border, or about 30 altogether (8). The line migrates ventrally to reach its final position at the juvenile stage, and an axonal branch follows each neuromast during this migration.Axonal regeneration after nerve cut has been demonstrated in the PLL during the first few days of life, leading to complete reinnervation of neuromasts within 24 h (9, 10). Regenerating axons follow either the Schwann cells that ensheathe the nerve (10) or the interneuromast cells that extend between consecutive neuromasts (11). PLL nerve regeneration has not yet been addressed beyond the early larval stage.Here we examine whether the PLL nerve is able to regenerate in adult and aging zebrafish. Our data clearly demonstrate the effectiveness and fidelity of regeneration at all ages studied, from 1 to 15 mpf, although the onset of reinnervation is increasingly delayed with age, thereby linking neuronal aging with a progressive decline in neuronal reactivity to axonal damage. Whenever a second cut is made after complete regeneration, the latency of reinnervation is reduced at all ages, provided the second cut is immediately distal to the first one. We show that although Schwann cells act as guidance cues to help the axons regrow along their original path, they are not involved in this regeneration-promoting effect. We conclude that the promoting effect of a first lesion is mostly caused by an intrinsic, local change occurring in the injured axons. Altogether, our results reveal that the zebrafish PLL is a convenient system to study axonal regeneration in vivo compared with mammalian systems, in which nerve wiring is more complex and less traceable.     

Risk Factors for Diarrhea in Children under Five Years of Age Residing in Peri-urban Communities in Cochabamba,Bolivia

This study examined the relationship between childhood diarrhea prevalence and caregiver knowledge of the causes and prevention of diarrhea in a prospective cohort of 952 children < 5 years of age in Cochabamba, Bolivia. The survey of caregiver knowledge found that more than 80% of caregivers were unaware that hand washing with soap could prevent childhood diarrhea. Furthermore, when asked how to keep food safe for children to eat only 17% of caregivers reported hand washing before cooking and feeding a child. Lack of caregiver awareness of the importance of practices related to hygiene and sanitation for diarrhea prevention were significant risk factors for diarrheal disease in this cohort. The knowledge findings from this study suggest that health promotion in these communities should put further emphasis on increasing knowledge of how water treatment, hand washing with soap, proper disposal of child feces, and food preparation relate to childhood diarrhea prevention.     

Clinical and laboratory characteristics of children with venous thromboembolism and protein C‐deficiency: an observational Israeli‐German cohort study

Venous thromboembolism [TE] is a multifactorial disease and protein C deficiency [PCD] constitutes a major risk factor. In the present study the prevalence of PCD and the clinical presentation at TE onset, including neonatal purpura fulminans, in a cohort of children are reported. In 367 unselected children (0·1–19 years) recruited between July 1996 and December 2013, a comprehensive thrombophilia screening was performed along with recording of anamnestic data. Twenty‐five of 338 children (7·4%) had PCD. Mean age at first TE onset was 10 years (range 0·1–18). Leading thromboembolic manifestations were neonatal purpura fulminans (n = 5), TE of cerebral veins (n = 3), stroke (n = 2) deep veinthrombosis (DVT) of the leg (n = 10), DVT & pulmonary embolism (n = 2) and DVT & pelvic veins (n = 3). Concomitant risk factors for TE were identified in 12 patients, whereas 13 children spontaneously developed TE. A positive family history of DVT was found in 10 children. In this unselected cohort of paediatric patients with symptomatic TE the overall prevalence of PCD was 7·4%; 1·5% presented with neonatal purpura fulminans. Given its clinical implication for patients and family members, thrombophilia testing should be performed and the benefit of medical or educational interventions should be evaluated in this high‐risk population.