Emanuel D. Pollack

Target-dependent survival of tadpole spinal cord neurites in tissue culture

Survival of nerve fibers growing from frog tadpole spinal cord explants in tissue culture was examined as a function of the absence or presence of target limb tissue. Neuritic outgrowth was maintained in serum-free medium for significantly longer times when cord was co-cultured with young limb tissue or limb conditioned medium than when grown alone. The results are suggested to be relevant to possible mechanisms controlling the loss of neurons from the spinal motor centers during in vivo development.

Stage dependency in eliciting target-dependent enhanced neurite outgrowth from spinal cord explants in vitro☆

Abstract Explants of frog tadpole spinal cord exhibit varying degrees of enhanced neuritic outgrowth in tissue culture that is dependent on the presence of appropriately staged limb target tissue. Nerve fiber outgrowth from early larval spinal cords was greatly augmented in the presence of early larval limb tissue, but was unresponsive to older limb tissues. Spinal cord explants of midlarval animals, on the other hand, failed to respond significantly to limb tissues over a broad developmental range. Nerve-target growth interactions must be regarded as developmental time-dependent systems with consideration of the differentiative state of both components.

Cyclic remodelling of growth cone lamellae and the effect of target tissue.

We report the existence of cyclical fluctuations in the total size of growth cone lamellae, represented by membrane protrusions and retractions, and show that aspects of this behavior can be regulated by the target tissue for the nerve fibers. The transition of the growth cone from a high to a less motile state, which occurs in the presence of the target tissue, has implications for the mechanisms that underlie nerve fiber elongation during development.

Late-generated cells in the lateral motor columns of developing frog spinal cord

Spinal motor neurons of larval frogs continue to be generated for the lateral motor columns (LMCs) even after their peak numbers have been reached and during the major period of normal neuronal loss. This study elucidates the extent to which relatively late generated motor neurons survive to the end of larval development. Bromodeoxyuridine was injected into Rana pipiens tadpoles at mid-larval stages twice a day for 5 days in order to label LMC neurons arising during the period of greatest neuron loss. Counts of labeled neurons visualized by immunocytochemical methods revealed that substantial numbers of late generated motor neurons survive to the end of larval development and may be retained in greater proportion than early generated motor neurons (prior to the onset of normally occurring neuron loss). Tadpoles that had been exposed to exogenous thyroxine so as to increase proliferative activity and thereby label more motor neurons during the injection period, exhibited an increase in labeled LMC cells although the total number of neurons remained comparable to controls. Protracted neurogenesis concurrent with cell loss during the later phases of development is a major determinant of the final population of the LMC.

Quantitative compensation by lateral motor column neurons in response to four functional hindlimbs in a frog tadpole

An analysis of the lateral motor columns of a frog tadpole with 4 functional unilateral hindlimbs revealed a 58% increase in the number of neurons on the overloaded side as compared to the control side of the spinal cord. An accompanying, apparently selective, increase in the number of very large neurons suggests that spinal motor centers respond to peripheral overload through a variable combination of neuronal number and size in providing for adequate innervation.

Growth rate analysis of neurites extended from triploid spinal cord explants.

Nerve fibers extending from spinal cord explants of triploid frog tadpoles were analyzed by time-lapse cinephotomicrography. The triploid neurites grew at a mean rate that seemed to reflect the increased chromosomal dosage and had maximal velocities that were several times greater than for diploid neurites. Concurrently, the time between alternating episodes of acceleration and deceleration in triploid neurite elongation phases was decreased. During a 24-h period, however, triploid neurites spent less time elongating with a net result of shorter travel distances than for diploid neurites. The data suggest that the genome is one level of nerve fiber growth control that can be manipulated by dosage alterations.

Stimulation of choline acetyltransferase in spinal cord explants by limb mesenchyme

Choline acetyltransferase (ChAT) activity in developing spinal cord explants in vitro is shown to be dependent on the presence of co‐cultured immature limb tissue. Frog tadpole spinal cord explants grown on collagen or polylysine expressed stage‐appropriate levels of ChAT activity only when in the presence of the limb mesenchyme target. Neither skeletal muscle nor polylysine, both of which enhance neurite growth accompanied by increases in cord protein, were capable of maintaining the level of ChAT activity characteristic of these spinal cords in vivo. The results demonstrate that developmentally significant levels of ChAT can be maintained in vitro under appropriate conditions that may act in part through the maintenance of cholinergic motor neurons.

Selective incorporation of 2-deoxyglucose into the developing spinal cord in vitro in response to target tissue and substratum

Differential uptake of [3H]deoxyglucose by larval frog spinal cord explants in vitro has demonstrated that metabolic activity is localized to the lateral motor columns in response to co-cultured target tissue or a neurite-promoting substratum. Autoradiographic data suggest that chemokinesis may be an integral component of motor neuron growth activation.