Alternative splicing: functional diversity among voltage-gated calcium channels and behavioral consequences.

Neuronal voltage-gated calcium channels generate rapid, transient intracellular calcium signals in response to membrane depolarization. Neuronal Ca(V) channels regulate a range of cellular functions and are implicated in a variety of neurological and psychiatric diseases including epilepsy, Parkinson's disease, chronic pain, schizophrenia, and bipolar disorder. Each mammalian Cacna1 gene has the potential to generate tens to thousands of Ca(V) channels by alternative pre-mRNA splicing, a process that adds fine granulation to the pool of Ca(V) channel structures and functions. The precise composition of Ca(V) channel splice isoform mRNAs expressed in each cell are controlled by cell-specific splicing factors. The activity of splicing factors are in turn regulated by molecules that encode various cellular features, including cell-type, activity, metabolic states, developmental state, and other factors. The cellular and behavioral consequences of individual sites of Ca(V) splice isoforms are being elucidated, as are the cell-specific splicing factors that control splice isoform selection. Altered patterns of alternative splicing of Ca(V) pre-mRNAs can alter behavior in subtle but measurable ways, with the potential to influence drug efficacy and disease severity. This article is part of a Special Issue entitled: Calcium channels.

Characterization of brainstem peptide YY (PYY) neurons.

Peptide YY (PYY), a member of the NPY superfamily of peptides, is predominantly synthesized by the colon and is thought to act on both the gut and brain to modulate energy homeostasis. Although neurons expressing PYY mRNA have also been reported in the brainstem, little is known about their physiological role and study of their projections has been problematic due to crossreactivity of PYY antibodies with NPY. In the present study we examined the localization of central PYY cell bodies in the mouse, rat, and monkey. In addition, efferent projections and afferent inputs of central PYY neurons were examined in rodents. Central PYY projections were examined by immunohistochemistry in the NPY knockout mouse, or with an NPY-preabsorbed PYY antibody in the rat to avoid any crossreactivity with NPY. In all species investigated PYY-immunoreactive (ir) cell bodies were localized exclusively to the gigantocellular reticular nucleus (Gi) of the rostral medulla. The highest density of PYY fibers was present within the solitary tract nucleus, specifically within the dorsal and lateral aspects. PYY fibers were also concentrated within the dorsal motor nucleus of the vagus and the hypoglossal nucleus. In addition, both orexin and melanin-concentrating hormone fibers made numerous close appositions with PYY cell bodies in the Gi. Collectively, the projection pattern and association with orexigenic neuropeptides suggest that brainstem PYY neurons may play a role in energy homeostasis through a coordinated effect on visceral, motor, and sympathetic output targets.

Cancer Survivors: The Success Story That's Straining Health Care.

Since President Richard Nixon declared a "War on Cancer" in 1971, the number of cancer survivors in the United States has quadrupled [1] and is still rising. Thanks to advance in cancer detection and treatment, the almost 15 million cancer survivors in the United States today could grow to some 19 million by 2024 [2]. Increasing survival rates have resulted in a shift: cancer is often treated as a chronic illness rather than a death sentence. However, having so many cancer survivors to monitor, track, and treat has led to growing pains for healthcare providers-forcing them to develop new ways to treat this increasing yet still vulnerable population.

Cell-Specific RNA Binding Protein Rbfox2 Regulates CaV2.2 mRNA Exon Composition and CaV2.2 Current Size.

The majority of multiexon mammalian genes contain alternatively spliced exons that have unique expression patterns in different cell populations and that have important cell functions. The expression profiles of alternative exons are controlled by cell-specific splicing factors that can promote exon inclusion or exon skipping but with few exceptions we do not know which specific splicing factors control the expression of alternatively spliced exons of known biological function. Many ion channel genes undergo extensive alternative splicing including Cacna1b that encodes the voltage-gated CaV2.2 α1 subunit. Alternatively spliced exon 18a in Cacna1b RNA encodes 21 amino acids in the II-III loop of CaV2.2, and its expression differs across the nervous system and over development. Genome-wide, protein-RNA binding analyses coupled to high-throughput RNA sequencing show that RNA binding Fox (Rbfox) proteins associate with CaV2.2 (Cacna1b) pre-mRNAs. Here, we link Rbfox2 to suppression of e18a. We show increased e18a inclusion in CaV2.2 mRNAs: (1) after siRNA knockdown of Rbfox2 in a neuronal cell line and (2) in RNA from sympathetic neurons of adult compared to early postnatal mice. By immunoprecipitation of Rbfox2-RNA complexes followed by qPCR, we demonstrate reduced Rbfox2 binding upstream of e18a in RNA from sympathetic neurons of adult compared to early postnatal mice. CaV2.2 currents in cell lines and in sympathetic neurons expressing only e18a-CaV2.2 are larger compared to currents from those expressing only Δ18a-CaV2.2. We conclude that Rbfox2 represses e18a inclusion during pre-mRNA splicing of CaV2.2, limiting the size of CaV2.2 currents early in development in certain neuronal populations.

Healing the Burn: Advances in Burn Treatment Technology Aim to Save Lives, Lessen Pain and Scarring.

When brothers Jamie and Glen Selby, aged 5 and 7, arrived at the Shriners Burns Institute in Denver, Colorado, in July 1983, more than 97% of their skin had been destroyed by a fire they had accidentally started while playing in an abandoned house. The boys were so badly burned that their outlook was grim-a 6-year-old friend who was also in the fire died from his injuries?but Jamie and Glen were lucky. Not only did they survive, but they were also some of the first patients to benefit from a new burn treatment nicknamed test-tube skin.

Orexin neurons express a functional pancreatic polypeptide Y4 receptor.

The receptor subtypes that mediate the effects of neuropeptide Y (NPY) on food intake have not been clearly defined. The NPY Y4 receptor has been identified recently as a potential mediator of the regulation of food intake. The purpose of the present study was to characterize the central site of action of the Y4 receptor using a combination of neuroanatomical and physiological approaches. Using immunocytochemistry, Y4-like immunoreactivity was found to be colocalized with orexin cell bodies in the lateral hypothalamic area (LHA) and orexin fibers throughout the brain. In situ hybridization confirmed the expression of Y4 mRNA in orexin neurons. To determine the functional interaction between Y4 receptors and orexin neurons, we examined the effects of rat pancreatic polypeptide (rPP), a Y4-selective ligand, or NPY, a nonselective ligand, administered directly into the LHA on the stimulation of food and water intake and c-Fos expression. Both rPP and NPY significantly increased food and water intake when they were administered into the LHA, although NPY was a more potent stimulator of food intake. Furthermore, both NPY and rPP significantly stimulated c-Fos expression in the LHA. However, whereas rPP stimulated c-Fos expression in orexin neurons, NPY did not. Neither rPP nor NPY stimulated c-Fos in melanin-concentrating hormone neurons, but both activated neurons of an unknown phenotype in the LHA. These results suggest that a functional Y4 receptor is expressed on orexin neurons and that these neurons are activated in response to a ligand with high affinity for the Y4 receptor (rPP). Although these data suggest a role for central Y4 receptors, the endogenous ligand for this receptor has yet to be clearly established.

Control of neuronal voltage-gated calcium ion channels from RNA to protein.

Voltage-gated calcium ion (CaV) channels convert neuronal activity into rapid intracellular calcium signals to trigger a myriad of cellular responses. Their involvement in major neurological and psychiatric diseases, and importance as therapeutic targets, has propelled interest in subcellular-specific mechanisms that align CaV channel activity to specific tasks. Here, we highlight recent studies that delineate mechanisms controlling the expression of CaV channels at the level of RNA and protein. We discuss the roles of RNA editing and alternative pre-mRNA splicing in generating CaV channel isoforms with activities specific to the demands of individual cells; the roles of ubiquitination and accessory proteins in regulating CaV channel expression; and the specific binding partners that contribute to both pre- and postsynaptic CaV channel function.

The neuronal splicing factor Nova controls alternative splicing in N-type and P-type CaV2 calcium channels.

Many cellular processes are involved in optimizing protein function for specific neuronal tasks; here we focus on alternative pre-mRNA splicing. Alternative pre-mRNA splicing gives cells the capacity to modify and selectively re-balance their existing pool of transcripts in a coordinated way across multiple mRNAs, thereby effecting relatively rapid and relatively stable changes in protein activity. Here we report on and discuss the coordinated regulation of two sites of alternative splicing, e24a and e31a, in P-type CaV2.1 and N-type CaV2.2 channels. These two exons encode 4 and 2 amino acids, respectively, in the extracellular linker regions between transmembrane spanning segments S3 and S4 in domains III and IV of each CaV2 subunit. Recent genome-wide screens of splicing factor-RNA binding events by Darnell and colleagues show that Nova-2 promotes inclusion of e24a in CaV2.2 mRNAs in brain. We review these studies and show that a homologous e24a is present in theCaV2 .1 gene, Cacna1a, and that it is expressed in different regions of the nervous system. Nova-2 enhances inclusion of e24a but represses e31a inclusion in CaV2.1 and CaV2.2 mRNAs in brain. It is likely that coordinated alternative pre-mRNA splicing across related CaV2 genes by common splicing factors, allows neurons to orchestrate changes in synaptic protein function while maintaining a balanced and functioning system.