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The present disclosure provides a method of testing or determining in vivo acute neurotoxicity of a molecule comprising measuring oscillations in intracellular free calcium concentration («calcium oscillations») in vitro in neuronal cells which are in contact with the molecule. E45. The method of any one of embodiments 1 to 44, further comprising measuring tubulin intensity of the molecule in a culture of neuronal cells. The above methods of the present invention can further comprise measuring tubulin intensity in a culture of neuronal cells, expression of a target protein, or behavioral performance of the molecule. In certain embodiments, the molecule comprises a protein, a peptide, a small molecule, a polynucleotide (e.g., an antisense oligonucleotide), or any combination thereof. E69. The method of any one of Embodiments 63 to 68, wherein the molecule comprises a small molecule, a polynucleotide, a protein, a peptide, or any combination thereof. E74. The method of any one of Embodiments 63 to 73, wherein the molecule is an antisense oligonucleotide. E56. The method of any one of embodiments 23 to 55, wherein the antisense oligonucleotide is a gapmer, a blockmer, a mixmer, or a wingmer. E87. In the method of any one of Embodiments 81 to 85, the improvement further comprising measuring an in vivo tolerability of the molecule
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11. The method of claim 1, further comprising measuring an in vivo tolerability of the molecule. In vivo tolerability scores were calculated based upon observations following a single intra-cerebroventricular (i.c.v.) administration of 100 .mu.g oligomers in mice or intra-thecal (i.t.) administration of 900 .mu.g oligomers or up to 1500 .mu.g in rats. E33. The method of any one of embodiments 1 to 32, further comprising measuring an in vivo tolerability of the molecule. E37. The method of any one of embodiments 34 to 36, wherein the tolerability category comprises at least one, at least two, at least three, at least four, or at least five tolerability categories. E73. The method of any one of Embodiments 63 to 72, further comprising measuring tublin intensity in a culture of neuronal cells. E14. The method of one of embodiments 1 to 13, further comprising administering the molecule to a subject in need of treatment of a disease or condition. E30. The method of any one of embodiments 23 to 29, wherein the antisense oligonucleotide modulates protein expression encoded by the target protein in the culture of the neuronal cells
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