Intramuscular administration of Botulinum toxin (BTx) has been associated with impaired osteogenesis in diverse conditions of bone formation (e.g., development, growth, and healing), yet the mechanisms of neuromuscular-bone crosstalk underlying these deficits have yet to be identified. Motivated by the emerging utility of zebrafish (Danio rerio) as a rapid, genetically tractable, and optically transparent model for human pathologies (as well as the potential to interrogate neuromuscular-mediated bone disorders in a simple model that bridges in vitro and more complex in vivo model systems), in this study we developed a model of BTx-induced muscle paralysis in adult zebrafish, and examined its effects on intramembranous ossification during tail fin regeneration. BTx administration induced rapid muscle paralysis in adult zebrafish in a manner that was dose-dependent, transient, and focal, mirroring the paralytic phenotype observed in animal and human studies. During fin regeneration, BTx impaired continued bone ray outgrowth, morphology, and patterning, indicating defects in early osteogenesis. Further, BTx significantly decreased mineralizing activity and crystalline mineral accumulation, suggesting delayed late-stage osteoblast differentiation and/or altered secondary bone apposition. Bone ray transection proximal to the amputation site focally inhibited bone outgrowth in the affected ray, implicating intra- and/or inter-ray nerves in this process. Taken together, these studies demonstrate the potential to interrogate pathological features of BTx-induced osteoanabolic dysfunction in the regenerating zebrafish fin, define the technological toolbox for detecting bone growth and mineralization deficits in this process, and suggest that pathways mediating neuromuscular regulation of osteogenesis may be conserved beyond established mammalian models of bone anabolic disorders.

Developmental dysplasia of the hip (DDH) is common because it is present in 1 of 100 newborns. Failure to diagnose DDH and treat in infancy can result in significant long-term disability. Early diagnosis can be accomplished through a quick but careful physical examination of all newborns. Further selective screening by ultrasound is indicated for those children with risk factors for DDH, which include family history, breech presentation, and unstable hip examination at the initial newborn examination. Continued examination of the hip at all routine well-child checkups is mandatory through the first year of life because late presenting DDH may occur. Treatment with a Pavlik harness is not typically instituted in the neonate because many unstable hips stabilize without intervention, but it is indicated in children older than 2 weeks with hip instability. Ultrasound screening for infants with risk factors for DDH is recommended at age 6 weeks. Pavlik harness treatment for children with unstable hips or significant dysplasia on ultrasound is continued until the hips stabilize and show concentric reduction on imaging. With time, diagnosis and treatment evolve to accommodate the growing child. Infants who fail to respond to nonoperative management may require more extensive interventions. At any time when treatment is initiated, a DDH specialist should be involved in the patient's care. If DDH is recognized early, treatment is less invasive, and long-term effects are minimized.