In vitro studies of spinal motoneuron development

Thesis/Dissertation · Tue Jan 01 04:00:00 EST 1985

OSTI ID:6942846

Calof, A L

Spinal motoneurons from chick embryos were purified by fluorescence-activated cell sorting after having been labelled by retrograde axonal transport of a lectin-fluorochrome conjugate. When these motoneurons were plated onto polylysine-coated dishes that had been exposed to medium conditioned over cultures of embryonic chick myotubes (MCM), they rapidly extended neurites. The neurite outgrowth-promoting activity in MCM was initially characterized by enzymatic digestions and sedimentation in associative cesium chloride gradients: These experiments suggested that both protein and heparan sulfate were important in the activity of the unfractionated conditioned medium. To further characterize the neurite outgrowth-promoting activity, MCM was metabolically labelled with /sup 35/S-methionine, and the activity was partially purified by salt precipitation and ion exchange chromatography and fractionated on Sepharose CL-4B. The peak of neurite outgrowth-promoting activity corresponded to the presence, in nonreducing SDS-polyacrylamide gels, of a protein band that comigrated with a laminin standard. Purified fractions of two putative neuronal trophic factors were tested for their ability to enhance motoneuron survival in culture. A 56,000 Da protein, purified on the basis of its binding to antibodies that block motor nerve terminal sprouting, had no effect on motoneuron survival. The Ciliary Neuronotrophic Factor (CNTF), a 20,000 Da protein which supports the survival of ciliary ganglion neurons in culture, enhanced motoneuron survival to some extent. Further experiments are needed to determine whether CNTF is responsible for some or all of the motoneuron survival-enhancing activity in MCM.

• Research Organization: California Univ., San Francisco (USA)
• OSTI ID: 6942846
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACIDS
ANIMAL CELLS
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIRDS
CARBOXYLIC ACIDS
CELL CULTURES
CELL DIFFERENTIATION
CELL FLOW SYSTEMS
CENTRAL NERVOUS SYSTEM
CHICKENS
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DRUGS
ELECTROPHORESIS
EVEN-ODD NUCLEI
FLUORESCENCE
FOWL
GEL PERMEATION CHROMATOGRAPHY
IN VITRO
ION EXCHANGE CHROMATOGRAPHY
ISOTOPE APPLICATIONS
ISOTOPES
LABELLED COMPOUNDS
LABELLING
LIGHT NUCLEI
LIPOTROPIC FACTORS
LUMINESCENCE
METHIONINE
NERVE CELLS
NERVOUS SYSTEM
NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
RADIOISOTOPES
SEDIMENTATION
SEPARATION PROCESSES
SOMATIC CELLS
SPINAL CORD
SULFUR 35
SULFUR ISOTOPES
TRACER TECHNIQUES
VERTEBRATES