Expression and Function of Serotonin 2A and 2B Receptors in the Mammalian Respiratory Network

Neurons of the respiratory network in the lower brainstem express a variety of serotonin receptors (5-HTRs) that act primarily through adenylyl cyclase. However, there is one receptor family including 5-HT2A, 5-HT2B, and 5-HT2C receptors that are directed towards protein kinase C (PKC). In contrast to 5-HT2ARs, expression and function of 5-HT2BRs within the respiratory network are still unclear. 5-HT2BR utilizes a Gq-mediated signaling cascade involving calcium and leading to activation of phospholipase C and IP3/DAG pathways. Based on previous studies, this signal pathway appears to mediate excitatory actions on respiration. In the present study, we analyzed receptor expression in pontine and medullary regions of the respiratory network both at the transcriptional and translational level using quantitative RT-PCR and self-made as well as commercially available antibodies, respectively. In addition we measured effects of selective agonists and antagonists for 5-HT2ARs and 5-HT2BRs given intra-arterially on phrenic nerve discharges in juvenile rats using the perfused brainstem preparation. The drugs caused significant changes in discharge activity. Co-administration of both agonists revealed a dominance of the 5-HT2BR. Given the nature of the signaling pathways, we investigated whether intracellular calcium may explain effects observed in the respiratory network. Taken together, the results of this study suggest a significant role of both receptors in respiratory network modulation

Supportive and symptomatic management of amyotrophic lateral sclerosis

The main aims in the care of individuals with amyotrophic lateral sclerosis (ALS) are to minimize morbidity and maximize quality of life. Although no cure exists for ALS, supportive and symptomatic care provided by a specialist multidisciplinary team can improve survival. The basis for supportive management is shifting from expert consensus guidelines towards an evidence-based approach, which encourages the use of effective treatments and could reduce the risk of harm caused by ineffective or unsafe interventions. For example, respiratory support using noninvasive ventilation has been demonstrated to improve survival and quality of life, whereas evidence supporting other respiratory interventions is insufficient. Increasing evidence implicates a causal role for metabolic dysfunction in ALS, suggesting that optimizing nutrition could improve quality of life and survival. The high incidence of cognitive dysfunction and its impact on prognosis is increasingly recognized, although evidence for effective treatments is lacking. A variety of strategies are used to manage the other physical and psychological symptoms, the majority of which have yet to be thoroughly evaluated. The need for specialist palliative care throughout the disease is increasingly recognized. This Review describes the current approaches to symptomatic and supportive care in ALS and outlines the current guidance and evidence for these strategies

Effect of cholesterol on the dipole potential of lipid membranes

The membrane dipole potential, ψd, is an electrical potential difference with a value typically in the range 150 – 350 mV (positive in the membrane interior) which is located in the lipid headgroup region of the membrane, between the linkage of the hydrocarbon chains to the phospholipid glycerol backbone and the adjacent aqueous solution. At its physiological level in animal plasma membranes (up to 50 mol%), cholesterol makes a significant contribution to ψd of approximately 65 mV; the rest arising from other lipid components of the membrane, in particular phospholipids. Via its effect on ψd, cholesterol may modulate the activity of membrane proteins. This could occur through preferential stabilization of protein conformational states. Based on its effect on ψd, cholesterol would be expected to favour protein conformations associated with a small local hydrophobic membrane thickness. Via its membrane condensing effect, which also produces an increase in ψd, cholesterol could further modulate interactions of polybasic cytoplasmic extensions of membrane proteins, in particular P-type ATPases, with anionic lipid headgroups on the membrane surface, thus leading to enhanced conformational stabilization effects and changes to ion pumping activity.Australian Research Counci

Genome-wide Analyses Identify KIF5A as a Novel ALS Gene

To identify novel genes associated with ALS, we undertook two lines of investigation. We carried out a genome-wide association study comparing 20,806 ALS cases and 59,804 controls. Independently, we performed a rare variant burden analysis comparing 1,138 index familial ALS cases and 19,494 controls. Through both approaches, we identified kinesin family member 5A (KIF5A) as a novel gene associated with ALS. Interestingly, mutations predominantly in the N-terminal motor domain of KIF5A are causative for two neurodegenerative diseases: hereditary spastic paraplegia (SPG10) and Charcot-Marie-Tooth type 2 (CMT2). In contrast, ALS-associated mutations are primarily located at the C-terminal cargo-binding tail domain and patients harboring loss-of-function mutations displayed an extended survival relative to typical ALS cases. Taken together, these results broaden the phenotype spectrum resulting from mutations in KIF5A and strengthen the role of cytoskeletal defects in the pathogenesis of ALS.Peer reviewe

Dual-wavelength ratiometric fluorescence measurement of the membrane dipole potential.

The electrostatic potentials associated with cell membranes include the transmembrane potential (delta psi), the surface potential (psi s), and the dipole potential (psi D). psi D, which originates from oriented dipoles at the surface of the membrane, rises steeply just within the membrane to approximately 300 mV. Here we show that the potential-sensitive fluorescent dye 1-(3-sulfonatopropyl)-4-[beta[2-(di-n-octylamino)-6- naphthyl]vinyl]pyridinium betaine (di-8-ANEPPS) can be used to measure changes in the intramembrane dipole potential. Increasing the content of cholesterol and 6-ketocholestanol (KC), which are known to increase psi D in the bilayer, results in an increase in the ratio, R, of the dye fluorescence excited at 440 nm to that excited at 530 nm in a lipid vesicle suspension; increasing the content of phloretin, which lowers psi D, decreases R. Control experiments show that the ratio is insensitive to changes in the membrane's microviscosity. The lack of an isosbestic point in the fluorescence excitation and emission spectra of the dye at various concentrations of KC and phloretin argues against 1:1 chemical complexation between the dye and KC or phloretin. The macromolecular nonionic surfactant Pluronic F127 catalyzes the insertion of KC and phloretin into lipid vesicle and cell membranes, permitting convenient and controlled modulation of dipole potential. The sensitivity of R to psi D is 10-fold larger than to delta psi, whereas it is insensitive to changes in psi S. This can be understood in terms of the location of the dye chromophore with respect to the electric field profile associated with each of these potentials. These results suggest that the gradient in dipole potential occurs over a span s5 A, a short distance below the membrane-water interface. These approaches are easily adaptable to study the influence of dipole potentials on cell membrane physiology