Abstract
Neurogenesis is a complex process leading to the generation of neuronal networks and glial cell types from stem cells or intermediate progenitors. Mapping subcellular and molecular changes accompanying the switch from proliferation to differentiation is vital for developing therapeutic targets for neurological diseases. Neuronal (N-type) and glial (S-type) phenotypes within the SH-SY5Y neuroblastoma cell line have distinct differentiation responses to 9-cis-retinoic acid (9cRA). In both cell phenotypes, these were accompanied at the single cell level by an uncoupling of Ca2+ store release from store-operated Ca2+ entry (SOCE), mediated by changes in the expression of calcium release-activated calcium pore proteins. This remodelling of calcium signalling was moderated by the predominant cell phenotype within the population. N- and S-type cells differed markedly in their phenotypic stability after withdrawal of the differentiation inducer, with the phenotypic stability of S-type cells, both morphologically and with respect to SOCE properties, in marked contrast to the lability of the N-type phenotype. Furthermore, the SOCE response of I-type cells, a presumed precursor to both N- and S-type cells, varied markedly in different cell environments. These results demonstrate the unique biology of neuronal and glial derivatives of common precursors and suggest that direct or indirect interactions between cell types are vital components of neurogenesis that need to be considered in experimental models.
Highlights
Mapping subcellular and molecular changes that accompany the cellular switch from proliferation to differentiation is vital for developing therapeutic targets for neurological diseases
We have previously shown that 9-cis-retinoic acid (9cRA)-induced differentiation of SH-SY5Y neuroblastoma cell populations induces an uncoupling of store-operated Ca2+ entry (SOCE) from endoplasmic reticulum (ER) Ca2+ store release [6, 22] coinciding with STIM1 and Orai1 down-regulation [6]
SOCE activation and initial deactivation rates remained elevated in differentiating N-type cells regardless of population, and in differentiating S-type cells in N-type-enriched populations, whereas in S-typeenriched populations, SOCE activation and initial deactivation rates of differentiating S-type cells were more similar to proliferating cells (Fig. 5). Previous findings from this laboratory have shown that SOCE becomes down-regulated in mixed [22] and enriched [6] populations of 9cRA-differentiated SH-SY5Y neuroblastoma cells
Summary
Mapping subcellular and molecular changes that accompany the cellular switch from proliferation to differentiation is vital for developing therapeutic targets for neurological diseases. Many studies of neuronal differentiation in vitro have used human neuroblastoma cell lines as models of human disease, and the SH-SY5Y cell line has attracted particular use as a model for Parkinson’s disease [2]. Whilst it is possible, with special manipulation, to obtain homogenous populations of neuronal SH-SY5Y cells [3], this cell line is of particular interest because both neuronal and glial-like cells can be identified within SH-SY5Y cultures. The cell line represents a powerful approach for studying the processes of neuronal and glial lineage development from the putative IPCs or I-type SH-SY5Y cells which can be identified within SH-SY5Y cell populations. Substrate adherent or glial-like S-type cells (Fig. 1g) express vimentin [6], fibronectin and tyrosinase [4, 7], markers characteristic of non-neuronal stromal or glial
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