Nerve cells of adult and aged mice grown in a monolayer culture: age-associated changes in morphological and physiological properties of dorsal root ganglion cells in vitro.

Abstract

In order to differentiate age-associated changes in morphological and physiological properties of mammalian nerve cells, dorsal root ganglion (DRG) cells of aged mice (C57BL/6; 98-99 weeks old) were grown in a monolayer culture. Neurite outgrowth, changes in shape and size of their soma and functional properties of their plasma membranes were compared to those of tissue-cultured DRG cells from young adult mice (4-8 weeks old). Trigeminal root ganglion (TRG) cells of aged mice were also grown in a monolayer culture, and their in vitro growth was compared to that of the aged DRG cells. Nerve cells were dissociated from DRG (or TRG) by digestion with collagenase and by trituration and were grown on collagen-coated plastic dishes for more than 14 days. Growth of neurites and changes in the size and shape of the nerve cell soma were viewed under a phase-contrast microscope, and physiological properties of the plasma membrane were studied by conventional intracellular recordings with a glass microelectrode. Both adult and aged DRG cells grew neurites of various length and underwent changes in shape and size of their soma, which could be divided into 2 stages; early and late. In the early stage of tissue culture (0-60 h in vitro), nerve cells altered their shape from a spherical to a spindle-like form. This change was not associated with the reduction in cell size. In the late stage of the tissue culture (3-14 days and thereafter), the DRG reduced their cell size, while changes in shape remained small. Quantitative comparison of the adult and aged DRG nerve cells revealed the following 3 major differences between 2 cultures: the survival fraction of the aged DRG cells counted at 36-48 h in vitro was 1/4 to 1/10 of that of the adult DRG cells in 3 different tissue culture trials; reduction in the cell size occurred much earlier in the aged than in the adult nerve cells; the rate of reduction in size of the aged DRG cells was large in comparison with that of the adult DRG cells. No difference in neurite growth or in physiological properties (resting membrane potential, input resistance, input capacitance or capability of generating both Na and Ca spikes) was detected between the aged and adult nerve cells in tissue culture.