Somatostatin receptor scintigraphy in the management of cerebral malignant ectomesenchymoma: a case report.

A 10-year-old girl presented with a cerebral malignant ectomesenchymoma (MEM), a very unusual tumour with undifferentiated mesenchymal as well as ectodermal elements. Somatostatin receptor scintigraphy (SRS) was performed during the diagnostic workup. The recurrent residual tumour mass was exactly visualized with SRS, and was negative after successful treatment of the patient. The potential application of SRS in initial staging, follow-up and therapy planning in MEM is discussed. This is the first application of SRS in MEM.

Prion protein affects Ca2+-activated K+ currents in cerebellar purkinje cells.

The prion protein (PrPC) has a primary role in the pathogenesis of transmissible spongiform encephalopathies. Its physiological function is not known yet. Altered late afterhyperpolarization has been observed in hippocampal CA1 pyramidal cells of prion protein-deficient mice (Prnp(0/0) mice) presumably caused by a disruption of Ca2+-activated K+ currents. An alteration of these currents has been recently described in scrapie-infected animals, and loss of function of PrPC has been put forward as one possible pathophysiological mechanism in prion diseases. This work focuses on patch-clamp studies of Ca2+-activated K+ currents in cerebellar Purkinje cells in the slice preparation of Prnp(0/0) mice as well as of transgenic mice. A significant correlation between PrPC expression in Purkinje cells and the maximal amplitude of TEA-insensitive Ca2+-activated K+ currents was observed, with reduced current amplitudes in Prnp(0/0) mice and a rescue of the phenotype in transgenic mice where PrPC had been reintroduced. Further studies of the intracellular free calcium concentration revealed an alteration of the maximal increase of intracellular calcium concentration with depolarization in the Prnp(0/0) mouse Purkinje cells. These data provide strong evidence that Ca2+-activated K+ currents in Prnp(0/0) mice are reduced due to an alteration of intracellular calcium homeostasis.

The cerebellum-specific Munc13 isoform Munc13-3 regulates cerebellar synaptic transmission and motor learning in mice.

Munc13 proteins form a family of three, primarily brain-specific phorbol ester receptors (Munc13-1/2/3) in mammals. Munc13-1 is a component of presynaptic active zones in which it acts as an essential synaptic vesicle priming protein. In contrast to Munc13-1, which is present in most neurons throughout the rat and mouse CNS, Munc13-3 is almost exclusively expressed in the cerebellum. Munc13-3 mRNA is present in granule and Purkinje cells but absent from glia cells. Munc13-3 protein is localized to the synaptic neuropil of the cerebellar molecular layer but is not found in Purkinje cell dendrites, suggesting that Munc13-3, like Munc13-1, is a presynaptic protein at parallel fiber-Purkinje cell synapses. To examine the role of Munc13-3 in cerebellar physiology, we generated Munc13-3-deficient mutant mice. Munc13-3 deletion mutants exhibit increased paired-pulse facilitation at parallel fiber-Purkinje cell synapses. In addition, mutant mice display normal spontaneous motor activity but have an impaired ability to learn complex motor tasks. Our data demonstrate that Munc13-3 regulates synaptic transmission at parallel fiber-Purkinje cell synapses. We propose that Munc13-3 acts at a similar step of the synaptic vesicle cycle as does Munc13-1, albeit with less efficiency. In view of the present data and the well established vesicle priming function of Munc13-1, it is likely that Munc13-3-loss leads to a reduction in release probability at parallel fiber-Purkinje cell synapses by interfering with vesicle priming. This, in turn, would lead to increases in paired-pulse facilitation and could contribute to the observed deficit in motor learning.

Altered intracellular calcium homeostasis in cerebellar granule cells of prion protein-deficient mice.

Previous studies have indicated that recombinant cellular prion protein (PrP(C)), as well as a synthetic peptide of PrP(C), affects intracellular calcium homeostasis. To analyze whether calcium homeostasis in neurons is also affected by a loss of PrP(C), we performed microfluorometric calcium measurements on cultured cerebellar granule cells derived from prion protein-deficient (Prnp(0/0)) mice. The resting concentration of intracellular free calcium [Ca(2+)](i) was found to be slightly, but significantly, reduced in Prnp(0/0) mouse granule cell neurites. Moreover, we observed a highly significant reduction in the [Ca(2+)](i) increase after high potassium depolarization. Pharmacological studies further revealed that the L-type specific blocker nifedipine, which reduces the depolarization-induced [Ca(2+)](i) increase by 66% in wild-type granule cell somas, has no effect on [Ca(2+)](i) in Prnp(0/0) mouse granule cells. Patch-clamp measurements, however, did not reveal a reduced calcium influx through voltage-gated calcium channels in Prnp(0/0) mice. These data clearly indicate that loss of PrP(C) alters the intracellular calcium homeostasis of cultured cerebellar granule cells. There is no evidence, though, that this change is due to a direct alteration of voltage-gated calcium channels.

c-myc oncogene family expression in glioblastoma and survival.

BACKGROUND: In gliomas, c-myc proto-oncogene expression has been found to correlate with the grade of malignancy, with low expression in Grade I and II and high expression in Grade III and IV tumors. We aimed to discover if myc expression is of prognostic significance in glioblastomas. METHODS: Expression of the c-myc, N-myc, and L-myc proto-oncogenes and of the max gene was investigated in 46 supratentorial glioblastomas from adult patients using in situ hybridization. RESULTS: Seventy-eight percent of the tumors expressed c-myc m-RNA, 84% max m-RNA, 57% N-myc m-RNA, and 57% L-myc m-RNA. The postoperative survival of patients over 60 years of age and that of patients under 60 years of age were analyzed separately, since advancing age was found to be negatively correlated with the duration of postoperative survival (p = 0.004). There was no significant difference in postoperative survival in either age group between patients whose tumors expressed either c-myc, N-myc, or L-myc, respectively, and those whose tumors did not exhibit this characteristic. A difference in postoperative survival, however, was found in the over 60-year age group between patients whose tumors expressed max to an equal or lesser extent than c-myc and those whose tumors expressed max to a greater extent than c-myc or neither max nor c-myc. CONCLUSION: The biologic behavior of glioblastomas in older patients may depend on the relative, but not on the absolute content of the c-myc protein and interacting proteins.

Microglial expression of the prion protein.

The prion protein (PrPc) is a normal cellular protein expressed by neurones and astrocytes. An altered isoform, PrPSc is thought to transmit spongiform encephalopathies. Here we show that microglia also express PrPc. Sensitivity of microglia to activation is enhanced by increased expression of PrPc. Bacterial endotoxin increases superoxide production and inhibits profileration of microglia more effectively when microglia express PrPc. PrPc expression is therefore important for the normal function of microglia.

The cellular prion protein binds copper in vivo.

The normal cellular form of prion protein (PrPC) is a precursor to the pathogenic protease-resistant forms (PrPSc) believed to cause scrapie, bovine spongiform encephalopathy (BSE) and Creutzfeldt-Jakob disease. Its amino terminus contains the octapeptide PHGGGWGQ, which is repeated four times and is among the best-preserved regions of mammalian PrPC. Here we show that the amino-terminal domain of PrPC exhibits five to six sites that bind copper (Cu(II)) presented as a glycine chelate. At neutral pH, binding occurs with positive cooperativity, with binding affinity compatible with estimates for extracellular, labile copper. Two lines of independently derived PrPC gene-ablated (Prnp0/0) mice exhibit severe reductions in the copper content of membrane-enriched brain extracts and similar reductions in synaptosomal and endosome-enriched subcellular fractions. Prnp0/0 mice also have altered cellular phenotypes, including a reduction in the activity of copper/zinc superoxide dismutase and altered electrophysiological responses in the presence of excess copper. These findings indicate that PrPC can exist in a Cu-metalloprotein form in vivo.

Increase of intracellular free Ca2+ in microglia activated by prion protein fragment.

A synthetic peptide consisting of amino acid residues 106 to 126 of the human prion protein (PrPc) that forms fibrils in vitro is toxic to cultured neurons. We have previously shown that the neurotoxic effect of this peptide is related to microglia activation (Brown et al., 1996a). For closer insight into this process of activation, we investigated the effect of the peptide on the intracellular free Ca2+ concentration ([Ca2+]i) in cultured microglia using Fura-2. Cultured microglia from wild-type as well as from PrPc gene-ablated mice (Prn-p0/0) responded to exposure to PrP106-126 with an increase in intracellular free calcium within 30 min. We observed two types of responses. Both in wild-type and Prn-p0/0 mice about half of the tested cells presented a small and often transient calcium increase after peptide application which was found to be independent of the extracellular calcium concentration. However, a further 33% of wild-type cells showed a strong and often permanent calcium increase depending on the extracellular calcium concentration, which was only rarely observed in Prn-p0/0 cells. To determine whether the response depended on the activation state of the microglia, we also examined LPS-treated activated microglia. The character of the calcium response remained unchanged, but significantly fewer cells responded. Our findings demonstrate the earliest reaction of microglia to a PrP fragment known to date.

PrP and beta-amyloid fragments activate different neurotoxic mechanisms in cultured mouse cells.

Alzheimer's disease and prion diseases such as Creutzfeldt-Jakob disease are caused by as yet undefined metabolic disturbances of normal cellular proteins, the amyloid precursor protein and the prion protein (PrP). Synthetic fragments of both proteins, beta-amyloid 25-35 (betaA25-35) and PrP106-126, have been shown to be toxic to neurons in culture. Cell death in both cases occurs by apoptosis. Here we show that there are considerable differences in the mechanisms involved. Thus, PrP106-126 is not toxic to cortical cell cultures of PrP knockout mouse neurons whereas betaA25-35 is. The toxicity of both peptides involves Ca2+ uptake through voltage-sensitive Ca2+ channels but only PrP106-126 toxicity involves the activity of NMDA receptors. The toxicity of betaA25-35, but not PrP106-126, is attenuated by the action of forskolin. These results indicate that PrP106-126 and PA25-35 induce neuronal apoptosis through different mechanisms.

Potential prognostic value of C-erbB-2 expression in medulloblastomas in very young children.

PURPOSE: The expression of the c-erbB-2 oncogene was studied in childhood medulloblastoma to evaluate its prognostic value, which has been claimed previously. PATIENTS AND METHODS: Tumor material from 45 patients < 15 years old at diagnosis was studied using 3 monoclonal antibodies against the internal and external domains of the c-erbB-2 oncogene product. RESULTS: Six of the 45 (13%) tumor specimens were found to be positive. C-erbB-2 expression was found more often in patients < 3 years old at diagnosis (4 of 15 patients, 27%) than in older patients (2 of 30, 6.6%). During the follow-up period (5.8 +/- 2.8 years) all patients with c-erbB-2 expression died of disease (after 1.2 +/- 0.7 years). Kaplan-Meier estimation revealed a highly significant correlation of c-erbB-2 expression and survival (p = 0.002). A further study of the expression of synaptophysin and the glial fibrillary acidic protein (GFAP) in the 45 tumors revealed a negative correlation of the expression of c-erbB-2 and these proteins. CONCLUSION: C-erbB-2, which may be predominantly expressed by less differentiated tumors, was found to delineate a poorer prognostic subgroup, especially when diagnosed in patients < 3 years old.

Patch-clamp analysis of synaptic transmission to cerebellar purkinje cells of prion protein knockout mice.

The prion protein (PrP) plays a pivotal role in transmissible spongiform encephalopathies such as Creutzfeldt-Jakob disease in humans and scrapie and bovine spongiform encephalopathy in animals. Previous experiments have suggested that the normal cellular prion protein (PrPc) is involved in synaptic function in the hippocampus. Here, we utilized the controlled recording conditions of the patch-clamp technique to investigate the synaptic function of prion protein in cerebellar Purkinje cells. By performing whole-cell and outside-out patch-clamp experiments in thin slices, we investigated synaptic transmission in prion protein knockout mice (PrP-null) and control animals. In PrP-null mice, the kinetics of GABA- and glutamate receptor-mediated currents showed no significant deviation from those in control animals. In contrast to previous results in hippocampal neurons, our findings support the view that synaptic transmission is unimpaired in prion protein-deficient mice.