RESUMEN
Lumbar spinal cord explants, harvested from neonatal rat pups aged between postnatal day 0 (P0) and P6, were cultured for a period of 48 hrs in the chemically defined medium R(12) on a poly-ethylene-imine (PEI) and on poly-D-lysin (PDL) coated surface. The outgrowth outside the explant was quantified. Lumbar explants from the same rat and embedded in a collagen matrix, and cortical explants from a P0 rat were used as controls. Statistical analysis demonstrated a clear relation between age-at-explantation and the number of neurites in the corona surrounding the explant. The number of outgrowing neurites decreased sharply with age-at-explantation. The average number of neurites per explant obeyed to the expression log (n) = -0.736x + 3.294 on PEI, and log (n) = -0.721x + 2.295 on PDL; x epsilon in [P0 - P6] (n, the number of neurites per explant; x, the age-at-explantation expressed in postnatal days). A similar observed age-related decrease of outgrowth has been described when culturing the lumbar explant inside a collagen matrix. The phenomenon appears to be an intrinsic property of the explant. We review growth inhibitory properties in different models and propose that the phenomenon occurs here at the interface explant-world.
Asunto(s)
Axones/fisiología , Técnicas de Cultivo de Órganos/métodos , Médula Espinal/fisiología , Animales , Colágeno/metabolismo , Ratas , Ratas Wistar , Factores de TiempoRESUMEN
Lumbar spinal cord explants, harvested from neonatal rat pups aged between postnatal day 0 (P0) and P7, were cultured for a period of 48 h in the chemically defined medium R(12) [17] (Romijn, H.J., van-Huijen, F., Wolters, P.S., Neurosci Biobehav Rev, 8 (1984) 301-334), embedded in a collagen matrix. The outgrowth into the surrounding matrix was quantified. Age-matched cortical explants were used as controls. Despite adaptations of the culture protocol, outgrowth remained variable. Statistical analysis demonstrated a clear relation between the age of the explant (at the time of explantation) and the number of neurites in the corona surrounding the explant. The number of outgrowing neurites decreased sharply with age. The average number of neurites per explant obeyed to the expression log(N)= -0.652 A+17 (N: the number of neurites per explant; A: the age expressed in gestational days; A epsilon [G23-G30]; G23 signifying gestational day 23, or P0). The observed age-related decrease of outgrowth could not be explained by progressive myelination of the spinal cord white matter, nor by the absence of trophic support from muscle, but may be related to a progressive inability of the spinal neurites to interact with collagen.
Asunto(s)
Técnicas de Cultivo de Célula/métodos , Neuronas Motoras/ultraestructura , Médula Espinal/citología , Factores de Edad , Animales , Animales Recién Nacidos , División Celular/efectos de los fármacos , Células Cultivadas , Colágeno/farmacología , Medios de Cultivo/farmacología , Femenino , Integrinas/metabolismo , Vértebras Lumbares , Masculino , Neuritas/metabolismo , Ratas , Ratas Wistar , Análisis de RegresiónRESUMEN
The axons that originate in the medial somatomotor cortex of the rat depart, during development, after those from the lateral somatomotor cortex, yet they arrive in the cervical spinal cord first. Either the medially originating axons elongate faster, or the laterally originating ones pause along the descent pathway. To investigate the presence of an intrinsic difference of the axonal elongation velocity between the lateral and medial somatomotor cortical areas, we cultured explants taken from these areas for 2 days, and measured the length of the outgrowth. After 2 days the explants were surrounded by a radiate corona of axons of which the longest measured 1.95 mm. A significant difference was detected between the medial and lateral somatomotor cortical areas in vitro. Axons originating from explants taken from the medial somatomotor cortical area are, after 2 days in culture, on average 0.16 mm longer than those from the lateral somatomotor cortical area. Though the observed difference is not large enough to allow for the overtaking observed in vivo, it does indicate that intrinsic differences exist within the developing rat somatomotor cortex. This in turn indicates that intrinsic cortical traits not only influence regionalization and targeting behavior of cortical projection neurons, but also their axonal elongation speed.