Adaptor protein complexes AP-4 and AP-5: new players in endosomal trafficking and progressive spastic paraplegia.

The adaptor proteins (APs) are a family of five heterotetrameric complexes with important functions in vesicle trafficking. While the roles of APs 1-3 are broadly established, comparatively little is known about AP-4 and AP-5. Current evidence suggests that AP-4 mediates TGN to endosome transport of specific cargo proteins, such as the amyloid precursor protein APP, and that it is involved in basolateral sorting in polarized cells. Furthermore, several independent studies have reported human patients with mutations in AP-4 genes. AP-4 deficiency causes severe intellectual disability and progressive spastic para- or tetraplegia, supporting an important role for AP-4 in brain function and development. The newly discovered AP-5 complex appears to be involved in endosomal dynamics; its precise localization and function are still unclear. Intriguingly, AP-5 deficiency is also associated with progressive spastic paraplegia, suggesting that AP-5, like AP-4, plays a fundamental role in neuronal development and homeostasis. The unexpected phenotypic parallels between AP-4 and AP-5 patients may in turn suggest a functional relationship of the two APs in vesicle trafficking.

Notch signalling stabilises boundary formation at the midbrain-hindbrain organiser.

The midbrain-hindbrain interface gives rise to a boundary of particular importance in CNS development as it forms a local signalling centre, the proper functioning of which is essential for the formation of tectum and cerebellum. Positioning of the mid-hindbrain boundary (MHB) within the neuroepithelium is dependent on the interface of Otx2 and Gbx2 expression domains, yet in the absence of either or both of these genes, organiser genes are still expressed, suggesting that other, as yet unknown mechanisms are also involved in MHB establishment. Here, we present evidence for a role for Notch signalling in stabilising cell lineage restriction and regulating organiser gene expression at the MHB. Experimental interference with Notch signalling in the chick embryo disrupts MHB formation, including downregulation of the organiser signal Fgf8. Ectopic activation of Notch signalling in cells of the anterior hindbrain results in an exclusion of those cells from rhombomeres 1 and 2, and in a simultaneous clustering along the anterior and posterior boundaries of this area, suggesting that Notch signalling influences cell sorting. These cells ectopically express the boundary marker Fgf3. In agreement with a role for Notch signalling in cell sorting, anterior hindbrain cells with activated Notch signalling segregate from normal cells in an aggregation assay. Finally, misexpression of the Notch modulator Lfng or the Notch ligand Ser1 across the MHB leads to a shift in boundary position and loss of restriction of Fgf8 to the MHB. We propose that differential Notch signalling stabilises the MHB through regulating cell sorting and specifying boundary cell fate.

Lrrn1 is required for formation of the midbrain-hindbrain boundary and organiser through regulation of affinity differences between midbrain and hindbrain cells in chick.

The midbrain-hindbrain boundary (MHB) acts as an organiser/signalling centre to pattern tectal and cerebellar compartments. Cells in adjacent compartments must be distinct from each other for boundary formation to occur at the interface. Here we have identified the leucine-rich repeat (LRR) neuronal 1 (Lrrn1) protein as a key regulator of this process in chick. The Lrrn family is orthologous to the Drosophila tartan/capricious (trn/caps) family. Differential expression of trn/caps promotes an affinity difference and boundary formation between adjacent compartments in a number of contexts; for example, in the wing, leg and eye imaginal discs. Here we show that Lrrn1 is expressed in midbrain cells but not in anterior hindbrain cells. Lrrn1 is down-regulated in the anterior hindbrain by the organiser signalling molecule FGF8, thereby creating a differential affinity between these two compartments. Lrrn1 is required for the formation of MHB--loss of function leads to a loss of the morphological constriction and loss of Fgf8. Cells overexpressing Lrrn1 violate the boundary and result in a loss of cell restriction between midbrain and hindbrain compartments. Lrrn1 also regulates the glycosyltransferase Lunatic Fringe, a modulator of Notch signalling, maintaining its expression in midbrain cells which is instrumental in MHB boundary formation. Thus, Lrrn1 provides a link between cell affinity/compartment segregation, and cell signalling to specify boundary cell fate.

Sustained interactive Wnt and FGF signaling is required to maintain isthmic identity.

Fibroblast growth factor 8 (FGF8) is expressed at the mid-hindbrain boundary and is an important signal emanating from the isthmic organizer. Wnt1 is expressed in the caudal midbrain juxtaposed to Fgf8 expression and has been implicated in its regulation. In this study, we examine the requirement for continuous Wnt signaling in the maintenance of Fgf8 expression at the isthmus. We demonstrate that prior to HH10, ongoing Wnt signaling is required to maintain the normal pattern of isthmic Fgf8 expression in ovo. Similarly, in explant assays, sustained Wnt signaling is essential to maintain Fgf8 expression in rhombomere 1. The mechanism by which Wnt signaling regulates isthmic Fgf8 expression is likely to be a maintenance response rather than an inductive effect. Finally, we show that Wnt maintenance of Fgf8 expression is dependent upon positive feedback by FGF signaling itself, and that rhombomere 1 does not receive instructive cues from the posterior hindbrain. In summary, these findings establish that a sustained reciprocal interaction between Wnt and FGF signaling is essential to maintain isthmic identity.

A Nice development: The first joint meeting of the British and French Societies for Developmental Biology, 13-16th September, 2003, Nice, France.

Held this autumn on the beautiful Cote d'Azur, the first joint meeting of the BSDB and SFBD provided delegates with the perfect informal setting for discussion spanning a broad cross-section of Developmental Biology. Participants' interests were diverse, ranging from the implementation of genome-wide approaches aimed at identifying all the molecular components of cell proliferation, signalling, patterning, and morphogenesis, to those engaged in capturing mesmerising glimpses of the minute and intricate workings of the cell. The meeting considered a wide spectrum of model organisms, including the simple plant Arabidopsis, the invertebrates Dictyostelium, Caenorhabditis elegans, and Drosophila melanogaster, the ascidian Ciona intestinalis, and the vertebrates Xenopus, zebrafish, chick, and mouse. Such a diverse approach served to highlight both similarities and differences in the molecular mechanisms that govern embryonic development among different species. Here, we highlight a few aspects of the meeting that illustrate this point.

Establishing the trochlear motor axon trajectory: role of the isthmic organiser and Fgf8.

Formation of the trochlear nerve within the anterior hindbrain provides a model system to study a simple axonal projection within the vertebrate central nervous system. We show that trochlear motor neurons are born within the isthmic organiser and also immediately posterior to it in anterior rhombomere 1. Axons of the most anterior cells follow a dorsal projection, which circumnavigates the isthmus, while those of more posterior trochlear neurons project anterodorsally to enter the isthmus. Once within the isthmus, axons form large fascicles that extend to a dorsal exit point. We investigated the possibility that the projection of trochlear axons towards the isthmus and their subsequent growth within that tissue might depend upon chemoattraction. We demonstrate that both isthmic tissue and Fgf8 protein are attractants for trochlear axons in vitro, while ectopic Fgf8 causes turning of these axons away from their normal routes in vivo. Both inhibition of FGF receptor activation and inhibition of Fgf8 function in vitro affect formation of the trochlear projection within explants in a manner consistent with a guidance function of Fgf8 during trochlear axon navigation.

Intramuscular droperidol versus intramuscular dimenhydrinate for the treatment of acute peripheral vertigo in the emergency department: a randomized clinical trial.

OBJECTIVE: The emergency department (ED) treatment of acute peripheral vertigo (APV) has not been well studied. The purpose of this study was to determine the efficacy of intramuscular (IM) droperidol vs IM dimenhydrinate, in the treatment of ED patients with APV. METHODS: The study was a randomized, double-blinded clinical trial, performed at a suburban, teaching ED. A convenience sample of adult patients with symptoms and signs consistent with rigid diagnostic criteria for APV were randomized to one of two treatment groups. Patients more than 65 years of age were excluded to reduce the likelihood of diagnostic misclassification. Demographic and historical features were recorded on a standardized data form. Patients recorded their initial level (t0) of discomfort on a 10-centimeter (cm) visual analog scale (VAS). Treatment group 1 received 2.5 mg droperidol IM, while treatment group 2 received 50 mg dimenhydrinate IM. After 30 minutes (t30), patients again recorded the severity of their symptoms on the VAS. Chi-square, t-tests, and Mann-Whitney were used for statistical comparison as appropriate. All tests were two-tailed, with alpha set at 0.05. Primary outcome parameters were the mean change in VAS score from t0 to t30, and the percentage of patients in each treatment group who felt well enough to go home after t30 without further ED intervention. RESULTS: There were 20 patients in the droperidol group and 20 in the dimenhydrinate group. The two groups were similar with respect to mean age (40 +/- 13 years droperidol vs. 42 +/- 13 years dimenhydrinate; p = 0.6), female sex (60% vs. 50%; p = 0.7), and mean median duration of symptoms [3 (interquartile range 2-12) vs 9 (interquartile range 2-30) hours; p = 0.2]. Mean initial t0 VAS scores were 7.2 +/- 2.3 and 7.8 +/- 1.9 (p = 0.47). Both treatment groups had mean reductions in VAS scores at t30 of 3.3 [95% confidence interval (95% CI) = 2.3 to 4.3]. At t30, 42% of patients in the droperidol group and 45% of patients in the dimenhydrinate group felt well enough to go home without further ED intervention. CONCLUSIONS: The authors found no difference between the therapeutic efficacies of IM droperidol and dimenhydrinate for the treatment of acute peripheral vertigo.