These neuronal properties in RA differentiated M17 cells were evidenced by both light microscopic observations (Figure?1) and immunofluorescence staining (Physique?2B – D). differentiated cells. Results We studied the effects of trans-retinoic acid treatment on (a) some differentiation marker proteins, (b) types of voltage-gated calcium (Ca2+) channels and (c) Ca2+-dependent neurotransmitter ([3H] glycine) release in cultured BE(2)-M17 cells. Cells treated with 10 M effects of these chemicals have been extensively reviewed in recent years and the issues Obatoclax mesylate (GX15-070) pertaining to their use have also been discussed [1-5]. The in vitro systems have been developed and utilized not only to understand the mechanisms of toxicity at the molecular and cellular levels but also to screen potential neurotoxicants. Potentially toxic compounds would be candidates for testing. The objective of neurotoxicologic studies on cells and tissues is usually to characterize the cellular and molecular substrates and pathways that contribute to impaired behavior, altered function, or pathological changes in the whole animal following exposure to a toxicant . The two main types of cell culture systems used for neurological testing are (a) primary neuronal cell cultures dissociated from peripheral or central nervous system tissues and (b) clonal cell lines derived from tumors of neurological origin . Primary neuronal cultures retain morphological, neurochemical, and electrophysiological properties of neurons models can provide a well-controlled system in which to study many of the critical cellular processes of neuronal development including proliferation, differentiation, growth, and synaptogenesis. Furthermore, cultured cell lines allow subtle changes in cell number, morphology, and functions to be readily detected compared to approaches and provide reproducibility in test results as well as providing a reduction in time, cost, and animal use [2,7]. Neuroblastoma cells can be differentiated by treatment with chemical agents into distinct morphologic cell types. These differentiated cells may be of different types: (a) substrate-adherent (S), which resemble non-neuronal precursor cells; (b) a sympathoadrenal neuroblastic (N); or (c) intermediate (I), which share elements of both S and N types . Each of these cell types differs in their ability to induce a tumor. N-type cells are malignant, where as the S-type cells are not; however, the I-type cells show the greatest malignancy [10,11]. One common neuroblastoma cell type used for research is BE(2)-M17, commonly known and henceforth called M17, which is available from ATCC. M17 is usually a human neuroblastoma cell line cloned from the SK-N-Be(2) neuroblastoma cell line isolated from a 2 year old male (ATCC, Manassas, VA). M17 cells Obatoclax mesylate (GX15-070) are multipotential with regard to neuronal enzyme expression e.g., choline acetyltransferase, acetylcholinesterase and dopamine–hydroxylase implying cholinergic, dopaminergic and adrenergic properties. M17 cells convert glutamate to GABA , however, this property is much less than that exhibited by BPTP3 cerebellar cortex which contains GABAergic neurons . There has been a great deal of research into differentiating the M17 cell line by treatment with effects of different neurotoxic substances [1,2,4,5]. Attempts have been made to develop and to utilize these in vitro neuronal models to study the mechanisms of toxicity due to chemical and biological compounds at cellular and molecular levels. Moreover, these models have also been tested for their use in rapid screening of potential neurotoxicants out of which positive compounds would be selected for evaluation. Prior studies using cellular models were intended Obatoclax mesylate (GX15-070) to generate preliminary mechanistic and toxicity information while reducing animal use and associated high cost of in vivo testing. The following are the three different types of cellular models primarily used in biomedical research; (1) primary cell cultures, (2) clonal cell lines, and (3) neural stem cells. The main advantage of using primary cell cultures is usually that they retain the morphological, neurochemical, and electrophysiological properties of neurons models: easy to obtain; relatively easy to grow; divide rapidly; and can be constantly subcultured for a relatively high number of passages to provide a large number of cells in a short period of time . The clonal M17 neuroblastoma cell line used in this study has the characteristics described above as well as the ability to become differentiated into a neuroblastic (N) cell when cultured in the presence of RA for several days [11,14]. These properties make the M17 cell line a good cell model for mechanistic and neurotoxicity testing. However, the functional changes in M17 cells due to RA differentiation have not been thoroughly characterized. A very relevant question is why do we need a differentiated neuronal model for neurobiology studies. The answer is usually that most of the neuronal functions such as membrane excitability, ion channels, neurotransmitter release, endocyctotic and exocyctotic events etc. are characteristics of.