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Compounds and methods for increasing neurogenesis

a neurogenesis and compound technology, applied in the field of compound and method for increasing neurogenesis, can solve the problems of limited fetal tissue grafting, small factors known to affect neurogenesis in vivo, and donor tissu

Inactive Publication Date: 2006-04-13
NEURONOVA AB COMPANY REGISTRATION NO 556703 0118
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the grafting of fetal tissue is limited by ethical considerations and a lack of donor tissue.
Yet, the number of factors known to affect neurogenesis in vivo is small and their effects are adverse or limited.

Method used

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  • Compounds and methods for increasing neurogenesis
  • Compounds and methods for increasing neurogenesis
  • Compounds and methods for increasing neurogenesis

Examples

Experimental program
Comparison scheme
Effect test

example 1

Reagents

[0148] Chemicals for dissociation of tissue included trypsin, hyaluronidase, and DNase (all purchased from SIGMA). Medium (DMEM 4.5 mg / ml glucose, and DMEM / F12), B27 supplement, and trypsin / EDTA were purchased from GIBCO. All plastic ware was purchased from ComingCostar. EGF for cell cultures was purchased from BD Biosciences, and the ATP-SL kit was purchased from BioThema.

[0149] For the test substances, the library was purchased from Phoenix pharmaceuticals Inc., USA, variety Pack Peptide Library (#L-001). Compounds purchased from Sigma-Aldrich included forskolin (#F6886), rolipram (#R6520), n-6,2-o-dibutyryladenosine (#D0260), cholera toxin (#C8052), MECA (#A024), HE-NECA (#H8034), nor-Binaltorphimine (#N1771), and adrenocorticotropic hormone (#A0298).

example 2

Mouse Neurosphere Cultures

[0150] The anterior lateral wall of the lateral ventricle of 5-6 week old mice was enzymatically dissociated in 0.8 mg / ml hyaluronidase and 0.5 mg / ml trypsin in DMEM containing 4.5 mg / ml glucose and 80 units / ml DNase at 37° C. for 20 minutes. The cells were gently triturated and mixed with Neurosphere medium (DMEM / F12, B27 supplement, 12.5 mM HEPES pH7.4), 100 units / ml penicillin and 100 μg / ml streptomycin. After passing through a 70 μm strainer, the cells were pelleted at 200×g for 4 minutes. The supernatant was subsequently removed and the cells were resuspended in Neurosphere medium supplemented with 3 nM EGF. Cells were plated out in culture dishes and incubated at 37° C. Neurospheres were ready to be split at approximately 7 days after plating.

[0151] To split neurosphere cultures, neurospheres were collected by centrifugation at 200×g for 4 minutes. The neurospheres were resuspended in 0.5 ml trypsin / EDTA in HBSS (1×), incubated at 37° C. for 2 minut...

example 3

ATP-Assay

[0152] To determine proliferation, neurospheres were split and seeded in Neurosphere medium as single cells in 96-well plates, at 10,000 cells / well. The following experiment was performed in sets of four parallel experiments (i.e., performed in quadruplicate) such that the cells may be used for different assays. Substances to be tested were added and cells were incubated at 37° C. for 4 days. Cells were lysed with 0.1% Triton-X100 in Tris-EDTA buffer. Intracellular ATP was measured using an ATP-SL kit according to the manufacturer's instructions (BioThema, Sweden). Intracellular ATP was shown to correlate with cell number. For each experiment, wells were visually examined for signs of neurogenesis and counted to confirm the results of the assay. Results were repeatable and statistically significant.

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Abstract

The invention is directed to methods of promoting neurogenesis by contacting neuronal tissue with neurogenesis modulating agents. Novel methods for treating neurological disorders using neurogenesis modulating agents are disclosed.

Description

RELATED APPLICATIONS [0001] This application is a divisional of U.S. patent application Ser. No. 10 / 850,055, filed May 19, 2004, which is a continuation-in-part of U.S. patent application Ser. No. 10 / 718,071 filed Nov. 30, 2003, which claims the benefit of priority to U.S. Provisional Application Ser. No. 60 / 427,912, filed Nov. 20, 2002. All patents and patent applications cited are hereby incorporated by reference.FIELD OF THE INVENTION [0002] The invention is directed to in vitro and in vivo methods of modulating neurogenesis. Novel agents for increasing intracellular levels of cAMP, Ca2+ and for modulating neurogenesis are also provided. BACKGROUND OF THE INVENTION [0003] Neural stem cells (NSC) are a source for new neurons in the mammalian CNS. NSC are located within the ependymal and / or subventricular zone (SVZ) lining the lateral ventricle (Doetsch et al., 1999; Johansson et al., 1999b) and in the dentate gyrus of the hippocampal formation (Gage et al., 1998). Studies have rev...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A61K38/22A61K38/23A61K31/675A61K38/26
CPCA61K31/675A61K38/23A61K38/2278A61K35/30C12N5/0619C12N2501/01C12N2501/60
Inventor BERTILSSON, GORANERLANDSSON, RIKARDFRISEN, JONASHAEGERSTRAND, ANDERSHEIDRICH, JESSICAHELLSTROM, NINAHAGGBLAD, JOHANJANSSON, KATARINAKORTESMAA, JARKKOLINDQUIST, PERLUNDH, HANNAMCGUIRE, JACQUELINEMERCER, ALEXNYBERG, KARLOSSOINAK, AMINAPATRONE, CESARERONNHOLM, HARRIETWIKSTROM, LILIANZACHRISSON, OLOF
Owner NEURONOVA AB COMPANY REGISTRATION NO 556703 0118
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