Lack of effects of 5-HT3 antagonists on normal and morphine-attenuated sexual behaviours in female and male rats

Although 5-HT₁ and 5-HT₂ receptor activity is known to influence copulation, the effects of 5-HT₃ receptor-selective drugs on sexual activity have yet to be systematically studied. The following experiments investigated the effects of the 5-HT₃-selective antagonists MDL 72222, ondansetron and ICS 205-930 on female sexual behaviour; male rats were studied using ondansetron and granisetron. These compounds influenced neither male nor female copulatory behaviours, suggesting that 5-HT₃ receptors contribute little to the modulation of sexual activity. 5-HT₃ receptor antagonists block certain opioid-induced behaviours and opioids selectively inhibit sexual behaviours; therefore, the ability of ondansetron and ICS 205-930 to modify morphine-attenuated copulatory activity was also tested. While morphine inhibited copulation, 5-HT₃ antagonists failed to reverse the effects.     

Insulin-producing cells in the brain of adult Drosophila are regulated by the serotonin 5-HT1A receptor

Insulin signaling regulates lifespan, reproduction, metabolic homeostasis, and resistance to stress in the adult organism. In Drosophila, there are seven insulin-like peptides (DILP1–7). Three of these (DILP2, 3 and 5) are produced in median neurosecretory cells of the brain, designated IPCs. Previous work has suggested that production or release of DILPs in IPCs can be regulated by a factor secreted from the fat body as well as by neuronal GABA or short neuropeptide F. There is also evidence that serotonergic neurons may regulate IPCs. Here, we investigated mechanisms by which serotonin may regulate the IPCs. We show that the IPCs in adult flies express the 5-HT_1A, but not the 5-HT_1B or 5-HT₇ receptors, and that processes of serotonergic neurons impinge on the IPC branches. Knockdown of 5-HT_1A in IPCs by targeted RNA interference (RNAi) leads to increased sensitivity to heat, prolonged recovery after cold knockdown and decreased resistance to starvation. Lipid metabolism is also affected, but no effect on growth was seen. Furthermore, we show that DILP2-immunolevels in IPCs increase after 5-HT_1A knockdown; this is accentuated by starvation. Heterozygous 5-HT_1A mutant flies display the same phenotype in all assays, as seen after targeted 5-HT_1A RNAi, and flies fed the 5-HT_1A antagonist WAY100635 display reduced lifespan at starvation. Our findings suggest that serotonin acts on brain IPCs via the 5-HT_1A receptor, thereby affecting their activity and probably insulin signaling. Thus, we have identified a second inhibitory pathway regulating IPC activity in the Drosophila brain.     

Characterization of the 5-HT1A receptor of the honeybee (Apis mellifera) and involvement of serotonin in phototactic behavior

Serotonin plays a key role in modulating various physiological and behavioral processes in both protostomes and deuterostomes. The vast majority of serotonin receptors belong to the superfamily of G-protein-coupled receptors. We report the cloning of a cDNA from the honeybee (Am5-ht1A) sharing high similarity with members of the 5-HT₁ receptor class. Activation of Am5-HT_1A by serotonin inhibited the production of cAMP in a dose-dependent manner (EC₅₀ = 16.9 nM). Am5-HT_1A was highly expressed in brain regions known to be involved in visual information processing. Using in vivo pharmacology, we could demonstrate that Am5-HT_1A receptor ligands had a strong impact on the phototactic behavior of individual bees. The data presented here mark the first comprehensive study—from gene to behavior—of a 5-HT_1A receptor in the honeybee, paving the way for the eventual elucidation of additional roles of this receptor subtype in the physiology and behavior of this social insect.     

Differential effects of the serotonin receptors on cultured rat cerebral cortical neurons

Effects of serotonin (5-HT) on cerebral cortical neurons were examined by patch clamp techniques. 5-HT produced a variety of responses such as outward (19/73 patches/neurons), slow inward (15/73 patches/neurons), fast inward (8/73 patches/neurons), and mixed currents (initially fast inward deflection followed by an outward response: 2/73 patches/neurons), with a latency of 12 sec, 15 sec, 0 sec, and 0 sec respectively, at a holding potential of −60 mV in whole-cell patches. The fast inward currents were again evoked by a selective 5-HT3 receptor agonist, 1-(m-chlorophenyl)-biguanide hydrochloride (CPBG). In the cell-attached patch clamp configuration, 5-HT inside the patch pipette elicited single channel currents with slope conductances of 42 pS and 132 pS (4/42 patches/neurons). CPBG inside the patch pipette evoked inward single channel currents with a lower slope conductance of 41 pS (3/23 patches/neurons). In contrast, application of 5-HT or a 5-HT₂ receptor agonist, α-methyl-5-hydroxytryptamine-maleate, outside the patch pipette induced outward single channel currents with a major slope conductance of 140 pS (8/30 patches/neurons) or 135 pS (6/20 patches/neurons), respectively. These results indicate that the outward and fast inward currents may be mediated respectively by the 5-HT₂ receptor, which is coupled to a G-protein, and by the 5-HT₃ receptor, which contains the non-selective cation channel, and that the mixed type may be caused by both the 5-HT₂ and 5-HT₃ receptors. Received 27 September 1996; received after revision 4 November 1996; accepted 7 November 1996     

Serotonin-stimulated protein phosphorylation in aortic smooth muscle cells

Summary The effects of serotonin on the formation of inositol phosphates and protein phosphorylation were examined in cultured smooth muscle cells. Serotonin stimulated the formation of [³H]inositol monophosphate, [³H]inositol bisphosphate and [³H]inositol trisphosphate. This effect was prevented by 5-HT₂ specific antagonist, 6-methyl-1-(1-methylethyl)ergoline-8-carboxylic acid, 2-hydroxy-1-methylpropyl ester [Z]-2-butenedioate (LY53857). Serotonin stimulated the phosphorylation of many polypeptides, among which a 20 kDa polypeptide was the most prominent. The phosphorylation was also inhibited by LY53857. LY53857 alone produced no effects on protein phosphorylation. The 20 kDa polypeptides were also phosphorylated by the addition of 12-O-tetradecanoylphorbol-13-acetate. These results suggest that serotonin stimulates protein phosphorylation through 5-HT₂ receptors and possibly activates protein kinase C in intact vascular smooth muscle cells.Part of the data contained in this paper was presented at the 74th local meeting of the Japanese Society of Pharmacology at Kanagawa.     

Structure and dynamics of rotary V<Subscript>1</Subscript> motor

Rotary ATPases are unique rotary molecular motors that function as energy conversion machines. Among all known rotary ATPases, F₁-ATPase is the best characterized rotary molecular motor. There are many high-resolution crystal structures and the rotation dynamics have been investigated in detail by extensive single-molecule studies. In contrast, knowledge on the structure and rotation dynamics of V₁-ATPase, another rotary ATPase, has been limited. However, recent high-resolution structural studies and single-molecule studies on V₁-ATPase have provided new insights on how the catalytic sites in this molecular motor change its conformation during rotation driven by ATP hydrolysis. In this review, we summarize recent information on the structural features and rotary dynamics of V₁-ATPase revealed from structural and single-molecule approaches and discuss the possible chemomechanical coupling scheme of V₁-ATPase with a focus on differences between rotary molecular motors.     

Augmentation of natural antiganglioside IgM antibodies in lower motor neuron disease (LMND) and role of CD5+ B cells

IgM antibodies directed against neuronal gangliosides GM₁ , GM₂ , GD_1a , GD_1b and GT_1b occur in normal individuals and their level significantly decreases with age. Patients with lower motor neuron disease (LMND) produce high levels of these autoantibodies. AntiGM₁ IgM is selectively augmented. In these patients, the CD5+ (B1) and CD5− (B2) subsets of B cells are not distinct entities but range from those without detectable CD5 marker to those with high CD5+ expression. B1 B cells were sorted to homogeneity, but B2 B cell cannot be isolated to homogeneity because of the presence of B1 cells with low CD5 expression. In short term cultures both the subsets produced IgM antibodies, but the antibodies reacted better with desialylated GM₁ than with GM₁ . Cycloheximide (Cx) (0.35 mM) largely blocked IgM synthesis of the B1 B cells but inhibition of the B2 B cells was incomplete, possibly due to shedding of cytophilic antibodies as well as to the presence of B1 phenotype with loss of CD5 expression. CD5+ B cells may be involved in the production of antiglycolipid IgM. Received 9 June 1997; received after revision 21 July 1997; accepted 28 July 1997     

The effect of glutamate,cysteate and related amino acids on insect muscle

Summary Application of Glu, CySO₂ or CySO₃ to blowfly larvae caused paralysis and increased the membrane potential of larval muscle. The contents of Glu and CySO₃ in larval muscle containing motor nerve terminals were markedly decreased after the perfusion of high K⁺ saline solution.     

Techniques simultanées de mesures: CO2(C14O2) élimination urinaire et activité motrice chez la souris dans l'expérimentation métabolique des amphétamines

Summary An apparatus which permits simultaneous measuring of the expiratory CO₂, the amount of C¹⁴O₂ and O₂ in metabolic studies of labelled drugs, is described. The urine and faeces are collected in the metabolic cage of this apparatus. Furthermore, the motor activity of the animal can be measured and recorded.     

Caffeine as a psychomotor stimulant: mechanism of action

The popularity of caffeine as a psychoactive drug is due to its stimulant properties, which depend on its ability to reduce adenosine transmission in the brain. Adenosine A₁ and A_2A receptors are expressed in the basal ganglia, a group of structures involved in various aspects of motor control. Caffeine acts as an antagonist to both types of receptors. Increasing evidence indicates that the psychomotor stimulant effect of caffeine is generated by affecting a particular group of projection neurons located in the striatum, the main receiving area of the basal ganglia. These cells express high levels of adenosine A_2A receptors, which are involved in various intracellular processes, including the expression of immediate early genes and regulation of the dopamine- and cyclic AMP-regulated 32-kDa phosphoprotein DARPP-32. The present review focuses on the effects of caffeine on striatal signal transduction and on their involvement in caffeine-mediated motor stimulation.Received 8 July 2003; received after revision 7 September 2003; accepted 6 October 2003     

Un curare d'une Erythrine africaine

Summary An alcaloid C₁₆H₁₉NO₃ has been isolated fromErythrina tholloniana; the iodohydrate of this erythroidine has a melting point of 225°C and 239°C for its chlorhydrate. It has a powerful curare-like action on the frog or on its isolated sciatic-sartorius preparation; at a concentration of less than 1/1,000,000, a complete neuromuscular block is produced: the electrical stimulation of the motor nerve does not produce any contraction, but the muscle reacts by an end-plate potential having the same characteristics (shape, duration, possibility of summation) as the electrical waves produced in the same preparation curarized by ordinary curare or by quaternary ammonium derivatives. Decurarization by veratrine 1/200,000 is accompanied by the same electrical reactions as those which have been described in preparations treated by curare.On mammals, the alcaloid has little curariform activity; on the isolated phrenic-diaphragm preparation of the rat, incomplete block was produced at a concentration of 1/5000.     

Role of 5-hydroxytryptamine in prostaglandin E1-induced potentiation of hexobarbitone hypnosis in albino rats

Summary PGE₁ potentiated, while diclofenac, a prostaglandin synthesis inhibitor, antagonized hexobarbitone hypnosis in rats. PGE₁-induced potentiation of hexobarbitone sleep was inhibited by a 5HT synthesis inhibitor and by a 5HT receptor blocker, suggesting that this potentiation is 5HT mediated.Acknowledgment. The gift of the following drugs are gratefully acknowledged: PGE₁ (Dr.J. E. Pike, Upjohn), diclofenac (Ciba-Geigy), methysergide (Sandoz) and hexobarbitone (Bayer).     

Acute cold exposure increases the glucagon sensitivity of thermogenic metabolism in the rat

Summary Administration of glucagon to rats at 25°C had no effect upon their VO₂ while administration of noradrenaline or noradrenaline plus glucagon raised the VO₂-At 5°C, noradrenaline had no effect upon the cold-enhanced VO₂, while glucagon caused a rise of 13.7% implying increased glucagon sensitivity at 5°C. The glucagon-induced enhancement of VO₂ was abolished by concurrent administration of noradrenaline.     

Zinc antagonizes the effect of botulinum type A toxin at the mouse neuromuscular junction

Summary Zn²⁺ (10–100 M) elevated the frequency of miniature end-plate potentials (MEPPs) in the mouse diaphragm. The effect did not depend on external Ca²⁺. Botulinum type A toxin (BTX_A, 50 ng/ml) abolished MEPPs almost completely within 30 min. Zn²⁺ (100 M) restored MEPPs and increased their frequency after they had been abolished by BTX_A in Ca²⁺-free solutions. The antagonistic effect of Zn²⁺ in the Ca²⁺-free solution was reduced by exposing the diaphragm to the toxin in the Ca²⁺-free solutions containing high K⁺. Thus, the action of BTX_A is probably enhanced by depolarization of the motor nerve terminals.     

One lipid, multiple functions: how various pools of PI(4,5)P2 are created in the plasma membrane

Phosphatidylinositol 4,5-bisphosphate [PI(4,5)P₂] is a minor lipid of the inner leaflet of the plasma membrane that controls the activity of numerous proteins and serves as a source of second messengers. This multifunctionality of PI(4,5)P₂ relies on mechanisms ensuring transient appearance of PI(4,5)P₂ clusters in the plasma membrane. One such mechanism involves phosphorylation of PI(4)P to PI(4,5)P₂ by the type I phosphatidylinositol-4-phosphate 5-kinases (PIP5KI) at discrete membrane locations coupled with PI(4)P delivery/synthesis at the plasma membrane. Simultaneously, both PI(4)P and PI(4,5)P₂ participate in anchoring PIP5KI at the plasma membrane via electrostatic bonds. PIP5KI isoforms are also selectively recruited and activated at the plasma membrane by Rac1, talin, or AP-2 to generate PI(4,5)P₂ in ruffles and lamellipodia, focal contacts, and clathrin-coated pits. In addition, PI(4,5)P₂ can accumulate at sphingolipid/cholesterol-based rafts following activation of distinct membrane receptors or be sequestered in a reversible manner due to electrostatic constrains posed by proteins like MARCKS.     

Does an inhibitor of mitochondrial adenylate kinase also affect oxidative phosphorylation?

Summary Adenylate kinase activity of intact mitochondria is strongly inhibited by Ap₅A, i.e.p ¹,p ⁵-Di (adenosine-5-) pentaphosphate, whereas oxidative phosphorylation is not affected. Therefore, Ap₅A is a useful tool to distinguish between oxidative and non oxidative ATP generating reactions.Acknowledgment. The generous support of Prof. Dr.Walther Lamprecht is gratefully acknowledged. J. L. thanks the Stipendienfonds des Verbandes der Chemische Industrie for a scholarship.     

Enhancement of Fe absorption by Mn in rice roots (Oryza sativa L.)

Summary The rates of absorption of Fe by excized rice roots were measured in the absence and presence of different concentrations of MnSO₄. Fe absorption from 0.1 and 5 mM FeSO₄ was enhanced by MnSO₄ at concentrations above 0.1 and 5 mM, respectively.     

Regulation of class V myosin

Class V myosin (myosin-5) is a molecular motor that functions as an organelle transporter. The activation of myosin-5′s motor function has long been known to be associated with a transition from the folded conformation in the off-state to the extended conformation in the on-state, but only recently have we begun to understand the underlying mechanism. The globular tail domain (GTD) of myosin-5 has been identified as the inhibitory domain and has recently been shown to function as a dimer in regulating the motor function. The folded off-state of myosin-5 is stabilized by multiple intramolecular interactions, including head–GTD interactions, GTD–GTD interactions, and interactions between the GTD and the C-terminus of the first coiled-coil segment. Any cellular factor that affects these intramolecular interactions and thus the stability of the folded conformation of myosin-5 would be expected to regulate myosin-5 motor function. Both the adaptor proteins of myosin-5 and Ca²⁺ are potential regulators of myosin-5 motor function, because they can destabilize its folded conformation. A combination of these regulators provides a versatile scheme in regulating myosin-5 motor function in the cell.     

eIF2A,an initiator tRNA carrier refractory to eIF2α kinases,functions synergistically with eIF5B

The initiator tRNA (Met-tRNA _i ^Met ) at the P site of the small ribosomal subunit plays an important role in the recognition of an mRNA start codon. In bacteria, the initiator tRNA carrier, IF2, facilitates the positioning of Met-tRNA _i ^Met on the small ribosomal subunit. Eukarya contain the Met-tRNA _i ^Met carrier, eIF2 (unrelated to IF2), whose carrier activity is inhibited under stress conditions by the phosphorylation of its α-subunit by stress-activated eIF2α kinases. The stress-resistant initiator tRNA carrier, eIF2A, was recently uncovered and shown to load Met-tRNA _i ^Met on the 40S ribosomal subunit associated with a stress-resistant mRNA under stress conditions. Here, we report that eIF2A interacts and functionally cooperates with eIF5B (a homolog of IF2), and we describe the functional domains of eIF2A that are required for its binding of Met-tRNA _i ^Met , eIF5B, and a stress-resistant mRNA. The results indicate that the eukaryotic eIF5B–eIF2A complex functionally mimics the bacterial IF2 containing ribosome-, GTP-, and initiator tRNA-binding domains in a single polypeptide.     

Bidirectional motility of kinesin-5 motor proteins: structural determinants,cumulative functions and physiological roles

Mitotic kinesin-5 bipolar motor proteins perform essential functions in mitotic spindle dynamics by crosslinking and sliding antiparallel microtubules (MTs) apart within the mitotic spindle. Two recent studies have indicated that single molecules of Cin8, the Saccharomyces cerevisiae kinesin-5 homolog, are minus end-directed when moving on single MTs, yet switch directionality under certain experimental conditions (Gerson-Gurwitz et al., EMBO J 30:4942–4954, 2011; Roostalu et al., Science 332:94–99, 2011). This finding was unexpected since the Cin8 catalytic motor domain is located at the N-terminus of the protein, and such kinesins have been previously thought to be exclusively plus end-directed. In addition, the essential intracellular functions of kinesin-5 motors in separating spindle poles during mitosis can only be accomplished by plus end-directed motility during antiparallel sliding of the spindle MTs. Thus, the mechanism and possible physiological role of the minus end-directed motility of kinesin-5 motors remain unclear. Experimental and theoretical studies from several laboratories in recent years have identified additional kinesin-5 motors that are bidirectional, revealed structural determinants that regulate directionality, examined the possible mechanisms involved and have proposed physiological roles for the minus end-directed motility of kinesin-5 motors. Here, we summarize our current understanding of the remarkable ability of certain kinesin-5 motors to switch directionality when moving along MTs.