Molecular Basis of Orb2 Amyloidogenesis and Blockade of Memory Consolidation.

Amyloids are ordered protein aggregates that are typically associated with neurodegenerative diseases and cognitive impairment. By contrast, the amyloid-like state of the neuronal RNA binding protein Orb2 in Drosophila was recently implicated in memory consolidation, but it remains unclear what features of this functional amyloid-like protein give rise to such diametrically opposed behaviour. Here, using an array of biophysical, cell biological and behavioural assays we have characterized the structural features of Orb2 from the monomer to the amyloid state. Surprisingly, we find that Orb2 shares many structural traits with pathological amyloids, including the intermediate toxic oligomeric species, which can be sequestered in vivo in hetero-oligomers by pathological amyloids. However, unlike pathological amyloids, Orb2 rapidly forms amyloids and its toxic intermediates are extremely transient, indicating that kinetic parameters differentiate this functional amyloid from pathological amyloids. We also observed that a well-known anti-amyloidogenic peptide interferes with long-term memory in Drosophila. These results provide structural insights into how the amyloid-like state of the Orb2 protein can stabilize memory and be nontoxic. They also provide insight into how amyloid-based diseases may affect memory processes.

Molecular mechanisms that change synapse number.

Synapses are the functional units of the nervous system, and their number and protein composition undergo changes over a wide time scale. These synaptic changes manifest into differential behavioural outputs and, in turn, changes in the external conditions to the individual may elicit changes in synapses. We review here publications appeared during the last 10 years in which advances on molecular and cellular mechanisms for synapse changes have been reported. We focus on synaptic changes occurring in the time range of minutes to hours, mainly.

Orb2 as modulator of Brat and their role at the neuromuscular junction.

How synapses are built and dismantled is a central question in neurobiology. A wide range of proteins and processes from gene transcription to protein degradation are involved. Orb2 regulates mRNA translation depending on its monomeric or oligomeric state to modulate nervous system development and memory. Orb2 is expressed in Drosophila larval brain and neuromuscular junction (NMJ), Orb2 knockdown causes a reduction of synapse number and defects in neuronal morphology. Brain tumor (Brat) is an Orb2 target; it is expressed in larval brain related with cell growth and proliferation. Brat downregulation induces an increase in synapse number and abnormal growth of buttons and branches in neurons. In absence of Orb2, Brat is overexpressed suggesting that Orb2 is negatively regulating Brat mRNA translation. Orb2 or Brat control the expression of specific genes related to neuronal function. Orb2 is required for Liprin and Synaptobrevin transcription meanwhile Brat is required for Synaptobrevin and Synaptotagmin transcription. We present here evidences of a novel genetic mechanism to regulate synapse fine tuning during development and propose an equilibrium between Orb2 conformational state and nervous system formation.

The guanine-exchange factor Ric8a binds to the Ca²âº sensor NCS-1 to regulate synapse number and neurotransmitter release.

The conserved Ca(2+)-binding protein Frequenin (homolog of the mammalian NCS-1, neural calcium sensor) is involved in pathologies that result from abnormal synapse number and probability of neurotransmitter release per synapse. Both synaptic features are likely to be co-regulated but the intervening mechanisms remain poorly understood. We show here that Drosophila Ric8a (a homolog of mammalian synembryn, which is also known as Ric8a), a receptor-independent activator of G protein complexes, binds to Frq2 but not to the virtually identical homolog Frq1. Based on crystallographic data on Frq2 and site-directed mutagenesis on Frq1, the differential amino acids R94 and T138 account for this specificity. Human NCS-1 and Ric8a reproduce the binding and maintain the structural requirements at these key positions. Drosophila Ric8a and Gαs regulate synapse number and neurotransmitter release, and both are functionally linked to Frq2. Frq2 negatively regulates Ric8a to control synapse number. However, the regulation of neurotransmitter release by Ric8a is independent of Frq2 binding. Thus, the antagonistic regulation of these two synaptic properties shares a common pathway, Frq2-Ric8a-Gαs, which diverges downstream. These mechanisms expose the Frq2-Ric8a interacting surface as a potential pharmacological target for NCS-1-related diseases and provide key data towards the corresponding drug design.

Small heat shock proteins determine synapse number and neuronal activity during development.

Environmental changes cause stress, Reactive Oxygen Species and unfolded protein accumulation which hamper synaptic activity and trigger cell death. Heat shock proteins (HSPs) assist protein refolding to maintain proteostasis and cellular integrity. Mechanisms regulating the activity of HSPs include transcription factors and posttranslational modifications that ensure a rapid response. HSPs preserve synaptic function in the nervous system upon environmental insults or pathological factors and contribute to the coupling between environmental cues and neuron control of development. We have performed a biased screening in Drosophila melanogaster searching for synaptogenic modulators among HSPs during development. We explore the role of two small-HSPs (sHSPs), sHSP23 and sHSP26 in synaptogenesis and neuronal activity. Both sHSPs immunoprecipitate together and the equilibrium between both chaperones is required for neuronal development and activity. The molecular mechanism controlling HSP23 and HSP26 accumulation in neurons relies on a novel gene (CG1561), which we name Pinkman (pkm). We propose that sHSPs and Pkm are targets to modulate the impact of stress in neurons and to prevent synapse loss.

[Descriptive study of the attitudes of family caregivers to the use of physical restraints: preliminary results]. / Estudio descriptivo sobre la actitud de la familia ante el uso de restricciones físicas en mayores: resultados preliminares.

OBJECTIVE: To evaluate the attitudes of families caring for elderly dependents to the use of physical restraints and to identify the factors that could influence these attitudes. MATERIAL AND METHODS: A descriptive, cross sectional study was carried out in a consecutive sample of the population, using a questionnaire investigating the various aspects that influence attitudes to restraints. Responses were obtained from 50 family caregivers, of which seven were discarded due to lack of knowledge about restraints (N=43). The characteristics of the sample, frequencies, the main trends and dispersion measures were analyzed. RESULTS: Most of the family caregivers considered the use of restrains to be appropriate and 90.7% were willing to use them with a relative; 41.9% believed that their use was unavoidable and were unaware of alternatives, while 23.3% were unaware that physical restraints can cause physical and psychological problems. None of the family caregivers believed that this method of restraint was a form of abuse. CONCLUSIONS: Relatives had a positive attitude to physical restraints and considered their use appropriate. This finding could be related to their limited knowledge of alternatives and of the complications that can be caused by restraints. Training programmes on alternatives to these devices should be instigated.

The equilibrium between antagonistic signaling pathways determines the number of synapses in Drosophila.

The number of synapses is a major determinant of behavior and many neural diseases exhibit deviations in that number. However, how signaling pathways control this number is still poorly understood. Using the Drosophila larval neuromuscular junction, we show here a PI3K-dependent pathway for synaptogenesis which is functionally connected with other previously known elements including the Wit receptor, its ligand Gbb, and the MAPkinases cascade. Based on epistasis assays, we determined the functional hierarchy within the pathway. Wit seems to trigger signaling through PI3K, and Ras85D also contributes to the initiation of synaptogenesis. However, contrary to other signaling pathways, PI3K does not require Ras85D binding in the context of synaptogenesis. In addition to the MAPK cascade, Bsk/JNK undergoes regulation by Puc and Ras85D which results in a narrow range of activity of this kinase to determine normalcy of synapse number. The transcriptional readout of the synaptogenesis pathway involves the Fos/Jun complex and the repressor Cic. In addition, we identified an antagonistic pathway that uses the transcription factors Mad and Medea and the microRNA bantam to down-regulate key elements of the pro-synaptogenesis pathway. Like its counterpart, the anti-synaptogenesis signaling uses small GTPases and MAPKs including Ras64B, Ras-like-a, p38a and Licorne. Bantam downregulates the pro-synaptogenesis factors PI3K, Hiw, Ras85D and Bsk, but not AKT. AKT, however, can suppress Mad which, in conjunction with the reported suppression of Mad by Hiw, closes the mutual regulation between both pathways. Thus, the number of synapses seems to result from the balanced output from these two pathways.

Complicaciones postratamiento del prolapso apical: a propósito de un caso / Complications after the treatment of apical prolapse: A case report

Resumen ANTECEDENTES: El prolapso de órganos pélvicos es un problema que puede resolverse con una diversidad de técnicas quirúrgicas según su tipo y características personales de la paciente. CASO CLÍNICO: Paciente de 81 años, originaria de Zaragoza, España, con índice de masa corporal de 41 kg/m2, hipertensión moderada y arritmia cardiaca, en tratamiento con acenocumarol y antihipertensivos de manera crónica. Antecedentes ginecológicos: tres embarazos de término que finalizaron en partos espontáneos, del segundo nació un niño de 4200 g. La paciente acudió a consulta por rectocele, corregido mediante la colocación de un pesario de anillo. El prolapso se complicó con rectoenterocele, que precisó corrección quirúrgica. Como consecuencia de la técnica quirúrgica elegida y por tratarse de una complicación frecuente de la vía de acceso (hematoma de la cúpula vaginal), la paciente sufrió una apertura vaginal a través de la que se hernió contenido intestinal. Después de evaluar el caso se decidió efectuar una nueva corrección quirúrgica que permitiera solucionar, simultáneamente, la hernia a través de la pared vaginal y la recidiva del prolapso apical. Hoy día se encuentra con adecuada evolución (12 meses del procedimiento quirúrgico), asintomática y sin complicaciones aparentes. CONCLUSIÓN: La cirugía del prolapso apical es compleja, debido a su amplia variedad de técnicas quirúrgicas y alto índice de recidiva. Es necesario conocer las diferentes vías de acceso para ofrecer la mejor solución a las pacientes.

Abstract BACKGROUND: Pelvic organ prolapse is a pathology that offers a variety of surgical techniques depending on the type of prolapse and the characteristics of the patient. CLINICAL CASE: Patient of 81 years born in Zaragoza (Spain) with a body mass index of 41kg / m2. It presents moderate hypertension and cardiac arrhythmia in treatment with anticoagulants. Requires treatment with acenocoumarol and antihypertensive in a chronic manner. Among the gynecological antecedents, there are three full-term pregnancies that ended with spontaneous deliveries, the second of them with a birth weight of 4,200gr. In this case, we present an elderly patient who initially presented a rectocele corrected initially using a pessary of the ring. The prolapse evolved presenting a rectoenterocele that required surgical correction. As a consequence of the chosen surgical technique and a frequent complication of the vaginal approach, such as a vaginal cuff hematoma, the patient suffered a vaginal opening through which intestinal contents were herniated. After evaluating the case, a new surgical correction was required that would allow the simultaneous resolution of the hernia through the vaginal wall that presented and the recurrence of the apical prolapse. Today is the right evolution (12 months of the surgical event), asymptomatic and without apparent complications. CONCLUSION: Prolapse surgery is complex due to its wide variety of surgical techniques and its high rate of recurrence. It is necessary to be aware of the different approaches to be able to offer the best solutions to our patients.