Myelination in cerebellar slice cultures: Development of a system amenable to biochemical analysis

Notterpek, Lucia; Bullock, P. N.; Malek-Hedayat, Shahnaz; Fisher, Robin S.; Rome, Leonard H.

doi:10.1002/jnr.490360603

Cited by 61 publications

(47 citation statements)

References 49 publications

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“…Therefore, the comparative study of this microenvironment is of special interest to validate our in vitro system. In vitro as in vivo, myelination starts during the first postnatal week (Seil, 1972;Reynolds and Wilkin, 1988;Notterpeck et al, 1993). Thus, in our mature-like slices, as expected, most Purkinje cell axons are myelinated, mimicking the conditions of adult cerebellum.…”

Section: Cerebellar Slices As An In Vitro System To Study Purkinje Cesupporting

confidence: 74%

Purkinje Cell Survival and Axonal Regeneration Are Age Dependent: AnIn VitroStudy

1997

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Purkinje cells are among the most resistant neurons to axotomy and the most refractory to axonal regeneration. By using organotypic cultures, we have studied age-and environmentrelated factors implicated in Purkinje cell survival and axonal regeneration. Most Purkinje cells taken from 1-to 5-d-old rats, the period in which these neurons are engaged in intense synaptogenesis and dendritic remodeling, die 1 week after plating, whereas if cultured before or after this period, Purkinje cells survive, even in the absence of deep nuclear neurons, their postsynaptic targets. Cerebellar slices taken from 10-dold rats and kept in vitro for 1 week acquire a cellular composition resembling mature cerebellum. Their Purkinje cells are resistant to axotomy, but even when confronted with permissive environments (sciatic nerves or fetal cerebellar slices), their axons do not regenerate. In contrast, fetal rat and mouse Purkinje cells are able to regenerate their axons on mature cerebellar slices. This regeneration is massive, and the regrowing axons invade all cerebellar regions of the apposed mature slices, including white matter. These results show that Purkinje cell survival and axonal regeneration are age-related and independent from environmental constraints. Moreover, our observations suggest strongly that the onset of synaptogenesis of Purkinje cell axons could provide a signal to turn off their growth program and that, thereafter, permissive microenvironment alone is unable to reestablish such a program. Key words: axonal regeneration; neuronal survival; cerebellum; Purkinje cell maturation; cerebellar organotypic cultures; axonal growthMost adult neurons survive axotomy if the lesion occurs far from their cell bodies, whereas perinatal neurons often die even after far-away axotomy (for review, see Schwab and Bartholdi, 1996). Frequently, this perinatal cell death can be counteracted by neurotrophic factors or by embryonic tissue grafting (for review, see Schwab and Bartholdi, 1996). In addition, immature CNS neurons have a higher potential to regenerate their axons than adult CNS neurons (for review, see Schwab and Bartholdi, 1996). Nevertheless, if allowed to regrow into transplanted peripheral nerves, most adult CNS cut axons regenerate their lesioned processes (David and Aguayo, 1981;Aguayo, 1985). Furthermore, the application of blocking antibodies against CNS myelin inhibitory proteins favors regeneration of axotomized corticospinal axons after spinal cord injury (Schnell and Schwab, 1990;Schwab et al., 1993). These observations strongly support the hypothesis that environmental factors rather than the intrinsic inability of adult CNS axons to regenerate are the limiting factor in the failure of axonal regeneration.Adult Purkinje cells are among the most resistant neurons to axotomy and the most refractory to axonal regeneration: they do not regenerate their axons even in the presence of their embryonic targets (Rossi et al., 1995). These cells in addition are one of the rare categories of neurons whose axons hav...

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Section: Cerebellar Slices As An In Vitro System To Study Purkinje Cesupporting

confidence: 74%

Purkinje Cell Survival and Axonal Regeneration Are Age Dependent: AnIn VitroStudy

1997

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“…For slice cultures, cerebella were dissected from neonatal Wistar rat pups (Ͻ24 hr old) (Notterpek et al, 1993). Meninges were caref ully removed, and tissue was incubated in Liebovitz's medium (Life Technologies, Gaithersburg, MD).…”

Section: Cell and Tissue Preparationsmentioning

confidence: 99%

“…Isolated slices were plated into poly-D-lysine-coated four-well plates and incubated in DM EM-F12 media containing 10% heat-inactivated horse serum, 20% fetal bovine serum, glucose (4.1 mg /ml), and insulin (15.6 g /ml). Slices were cultured for 3 weeks in decreasing concentrations of fetal bovine serum according to the protocol of Notterpek et al (1993).…”

Section: Cell and Tissue Preparationsmentioning

confidence: 99%

Pituitary Adenylyl Cyclase-Activating Polypeptide Stimulates DNA Synthesis But Delays Maturation of Oligodendrocyte Progenitors

Lee¹,

Lelièvre²,

Zhao³

et al. 2001

J. Neurosci.

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The neuropeptide pituitary adenylyl cyclase-activating peptide (PACAP) and one of its receptors (PAC 1 ) are expressed in embryonic neural tube, where they appear to regulate neurogenesis and patterning. We now show that PAC 1 gene expression is also present in neonatal rats in the ventricular and subventricular zones and in the optic chiasm, areas that are rich in oligodendrocyte (OL) progenitors (OLP). Because actions of PACAP on OLP have not been reported, we examined the effects of PACAP on the proliferation of purified OLP in culture and on myelinogenesis in cerebellar slices. Northern analyses on total RNA from purified glial cell subtypes revealed an abundant 7 kb hybridizing transcript in OLP, which was confirmed to correspond to the PAC 1 receptor by reverse transcription-PCR. The presence of this receptor was also corroborated by radioligand binding and cAMP assay. In cultured OL, receptor density decreased during maturation but was partially counterbalanced by the appearance of sites that bound both PACAP and the related peptide vasoactive intestinal peptide. PACAP increased DNA synthesis in OLP cultures almost twofold and increased the bromodeoxyuridine-labeling index in O4-positive OLP. PACAP treatment also resulted in decreased sulfate incorporation into sulfatide in cultures of differentiating OL. The PACAP effect on sulfatide synthesis was fully reproduced in a cerebellar explant model. These findings indicate that PACAP may act at two stages during OL development to (1) stimulate proliferation and (2) delay maturation and/or myelinogenesis.

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“…Myelinating slice cultures of rodent CNS have proven useful for removing the influences of the systemic immune system on myelination (Birgbauer et al, 2004;Mi et al, 2009;Notterpek et al, 1993). Unfortunately, these models are primarily derived from developing tissue and their ability to effectively mimic in vivo demyelination/remyelination is unclear (Birgbauer et al, 2004).…”

Section: Alternative Approaches To Mouse Models Of Ms In Vitro Modelsmentioning

confidence: 99%

Restoring the balance between disease and repair in multiple sclerosis: insights from mouse models

Miller

Fyffe-Maricich

2010

Disease Models &Amp; Mechanisms

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Multiple sclerosis (MS) is considered an autoimmune-mediated demyelinating disease that targets the central nervous system (CNS). Despite considerable research efforts over multiple decades, our understanding of the basic biological processes that are targeted in the disease and the mechanisms of pathogenesis are poorly understood. Consequently, current therapies directed at controlling the progression of the disease are limited in their effectiveness. Historically, the primary focus of MS research has been to define the cellular and molecular basis of the immunological pathogenic mechanisms. Recently, however, it has become clear that long-term functional recovery in MS will require the development of strategies that facilitate myelin repair in lesion areas. The emerging evidence that the adult vertebrate CNS retains the capacity to regenerate neural cells that have been lost to disease or damage has provoked intensive research focused on defining the mechanisms of myelin repair. Unfortunately, the existing animal models of MS are poorly equipped to assess myelin repair, and new validated strategies to identify therapeutics targeted at promoting myelin repair are badly needed. This Commentary will review established murine models of MS, and discuss emerging technologies that promise to provide insights into the mechanisms of myelin repair.

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Myelination in cerebellar slice cultures: Development of a system amenable to biochemical analysis

Cited by 61 publications

References 49 publications

Purkinje Cell Survival and Axonal Regeneration Are Age Dependent: AnIn VitroStudy

Purkinje Cell Survival and Axonal Regeneration Are Age Dependent: AnIn VitroStudy

Pituitary Adenylyl Cyclase-Activating Polypeptide Stimulates DNA Synthesis But Delays Maturation of Oligodendrocyte Progenitors

Restoring the balance between disease and repair in multiple sclerosis: insights from mouse models

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