Gating Pore Currents in Sodium Channels.

Voltage-gated sodium channels belong to the superfamily of voltage-gated cation channels. Their structure is based on domains comprising a voltage sensor domain (S1-S4 segments) and a pore domain (S5-S6 segments). Mutations in positively charged residues of the S4 segments may allow protons or cations to pass directly through the gating pore constriction of the voltage sensor domain; these anomalous currents are referred to as gating pore or omega (ω) currents. In the skeletal muscle disorder hypokalemic periodic paralysis, and in arrhythmic dilated cardiomyopathy, inherited mutations of S4 arginine residues promote omega currents that have been shown to be a contributing factor in the pathogenesis of these sodium channel disorders. Characterization of gating pore currents in these channelopathies and with artificial mutations has been possible by measuring the voltage-dependence and selectivity of these leak currents. The basis of gating pore currents and the structural basis of S4 movement through the gating pore has also been studied extensively with molecular dynamics. These simulations have provided valuable insight into the nature of S4 translocation and the physical basis for the effects of mutations that promote permeation of protons or cations through the gating pore.

Breast lumps in service women.

INTRODUCTION: Breast cancer is uncommon in a young population but it does occur. 80% of breast cancer occurs after 50 yrs of age. This article uses current guidelines and evidence to advise military medical staff on how best to investigate and manage serving-age women presenting with breast symptoms. Male breast changes will be dealt with in a future article. DIFFERENTIAL DIAGNOSIS: Young females presenting with breast lumps are unlikely to have cancer. In order of frequency the causes are likely to be benign breast change; fibroadenoma; abscesses in 20-30 year olds; cysts in 30-40 year olds, and lastly cancer. The UK sees 48,000 new cases of breast cancer in women every year; breast cancer can also occur in men but is very rare. DIAGNOSIS AND MANAGEMENT: Management in the deployed, primary and secondary care settings are described. It may be reasonable in young women to wait and see if a lump resolves after the patient's next menstrual cycle before referring the patient. Once referred, current guidelines recommend that all patients are seen by a breast surgeon within two weeks. Within this group, a subgroup of patients with 'red-flag' lumps is identified who need to be referred urgently. The remaining patients have lumps that can be considered non-urgent: however, hospitals will still endeavour to see these patients within two weeks.. CONCLUSIONS: Breast cancer is more difficult to diagnose in the younger patient. In primary care, breast lumps are still simple to manage if the points in this article are followed. Anxious patients can be reassured that cancer is unlikely. However, cancer in this young age group is associated with worse outcomes than breast cancer in older patients.

Characterization of crustacean cardioactive peptide-like immunoreactivity in the horseshoe crab, Limulus polyphemus.

The localization of crustacean cardioactive peptide-like immunoreactivity in the horseshoe crab Limulus polyphemus was investigated with enzyme-linked immunosorbent assay and fluorescence microscopy. Immunoreactivity was quantified in the opisthosomal nervous system (67.7 +/- 11.4 ng/g), cardiac ganglion (45.0 +/- 10.3 ng/g), prosomal nervous system (28.5 +/- 6.6 ng/g), and midgut (24.6 +/- 6.7 ng/g). In the brain, immunoreactive somata were observed in ganglion cells of the central body, in the medullary group and within the ventral medial group. Clusters of immunoreactive cells were found in each of the circumesophageal, pedal ganglia, and in the opisthosomal, abdominal ganglia. In the periphery, immunoreactive varicose fibers were observed in branches of the intestinal nerves, and near longitudinal and circular muscle fibers of the midgut. Immunoreactivity was observed in the cardiac ganglion and myocardium of the neurogenic heart. Synthetic crustacean cardioactive peptide had slight excitatory effects on the cardiac rhythm at doses up to 10(-6) M. This peptide had excitatory effects on the midgut at nanomolar doses. Ventral nerve cord extracts were partially purified with reverse phase high performance liquid chromatography. Two regions of immunoreactivity were detected, one of which coeluted with the authentic peptide. The distribution of crustacean cardioactive peptide immunoreactivity is compared with other transmitter systems in the Limulus nervous system, and myotropic actions of this peptide are discussed with respect to peptidergic modulation of intestinal motility.

Characterization and distribution of FMRFamide immunoreactivity in the rat central nervous system.

FMRFamide immunoreactive material (irFMRFamide) was studied in rat brain and gastrointestinal tract. Highest irFMRFamide concentrations were found in tissues of the gastrointestinal tract and, in the brain, highest concentrations were found in the hippocampus, midbrain, brainstem and hypothalamus. High pressure liquid chromatographic characterization of irFMRFamide demonstrated that the immunoreactive material in brain, pancreas and duodenum was different from molluscan FMRFamide but it was also distinct from any known neuropeptide.

Presence and distribution of immunoreactive and bioactive FMRFamide-like peptides in the nervous system of the horseshoe crab, Limulus polyphemus.

FMRFamide immunoreactivity was detected in all regions of the Limulus nervous system, including the brain (6.5 +/- 0.6 pg FMRFamide/mg), cardiac ganglion (2.06 +/- 0.67 pg FMRFamide/mg), and ventral nerve cord (5.8 +/- 0.7 pg FMRFamide/mg). The distribution of immunoreactive FMRFamide (irFMRFamide) was mapped by immunofluorescence and the distribution corresponded to regional RIA data. A good proportion of the CNS and cardiac ganglion neuropile contained irFMRFamide, and fluorescent cell bodies were observed in several areas. High performance liquid chromatography (HPLC) was employed to separate and characterize the FMRFamide-like peptides from extracts of Limulus brains. HPLC fractions were analyzed using coincidental radioimmunoassay and bioassay (the radula protractor muscle of Busycon contrarium). There appear to be at least three FMRFamide-like peptides in the Limulus brain, including one similar to clam FMRFamide. FMRFamide acts on Limulus heart in a biphasic manner at relatively high concentrations (10(-5)M), but has no effect on the activity of the isolated ventral nerve cord. These data suggest that in Limulus FMRFamide-like peptides are acting as neurotransmitters, or neuromodulators.

Identification of proctolin in the central nervous system of the horseshoe crab, Limulus polyphemus.

A proctolin-like peptide was isolated from the prosomal CNS of the chelicerate arthropod, Limulus, and purified using size exclusion, ion exchange and high performance liquid chromatography. Coincident bioassay (cockroach hindgut) and radioimmunoassay were employed to identify fractions which contained proctolin-like material. Proctolin-like activity coeluted with synthetic proctolin with all three chromatographic techniques employed. When applied to either the Limulus heart or hindgut preparations, purified Limulus proctolin produced excitatory responses which were indistinguishable from those produced by the synthetic peptide. Purified samples of the Limulus proctolin-like peptide were subjected to Edman degradation and tandem mass spectrometry and the amino acid sequence of the Limulus peptide was determined to be identical to that of cockroach proctolin (H-Arg-Tyr-Leu-Pro-Thr-OH). The presence of proctolin in the Limulus CNS and its biological action on the isolated heart and hindgut suggest a physiological role for this peptide in the regulation of cardiac output and hindgut motility.

Carcinoma developing in the neo-oesophagus following gastroplasty.

Carcinoma developing after gastroplasty and fundoplication has been documented in only five cases. We report a further case occurring in a patient 4 years after initial surgery. The tumour originated in the newly created tubular extension. There was no evidence of a columnar lining in the true oesophagus prior to surgery. The development, technique and complications of gastroplasty and partial fundoplication are discussed with special mention of the diagnostic problems when a tumour develops in a patient who has undergone this procedure.

Excitatory actions of FMRFamide-related peptides (FaRPs) on the neurogenic Limulus heart.

The actions of FMRFamide-related peptides (FaRPs) on the neurogenic heart of the horseshoe crab, Limulus polyphemus, were investigated. Excitatory chronotropic effects were produced by application of TNRNFLRFamide, SDRNFLRFamide, GYNRS-FLRFamide, or pQDPFLRFamide to the intact heart preparation. Effects were dose-dependent with a threshold of 10(-9) M or less. TNRNFLRFamide and SDRNFLRFamide increased the burst rate of the isolated Limulus cardiac ganglion. Synthetic FaRPs produced inotropic excitation of the heartbeat as well. GYNRSFLRFamide, TNRNFLRFamide, SDRNFLRFamide, and pQDPFLRFamide increased heart contraction strength at a threshold dose of approximately 10(-8) M. TNRNFLRFamide and SDRNFLRFamide enhanced electrically evoked contractions of the Limulus myocardium, elicited contracture in some preparations, and increased the excitability of cardiac muscle fibers. The presence of cardioactive FaRPs in the Limulus central nervous system was suggested by reverse phase HPLC of acidified methanol extracts of Limulus nervous tissue. Four peaks of FaRP-like bioactivity were detected with the Busycon radula protractor muscle bioassay. These peaks also contained FaRP-like immunoreactivity. Two of these partially purified peaks produced excitatory chronotropic effects on the intact Limulus heart preparation similar to those produced by synthetic FaRPs.

Glutamate as a transmitter in the sensory pathway from prostomial lip to serotonergic Retzius neurons in the medicinal leech Hirudo.

The involvement of glutamate in putative ingestive sensory pathways affecting the excitability of serotonergic Retzius neurons (RZ) in the leech CNS was investigated with a pharmacological approach. Exposure of the prostomial lip to 150 mM NaCl and 1 mM arginine produced excitatory as well as inhibitory responses in RZ found in the reproductive segments, while only excitatory responses were elicted in standard midbody RZ. Antagonists of glutamatergic receptors of the kainate/quisqualate type effectively inhibited chemosensory dependent excitation of RZ. Antagonists of glutamatergic receptors of the N-methyl D-aspartate type were ineffective in this regard. Cephalic nerve stimulation, like chemical stimulation of the lip, produced segment-specific responses in midbody RZ. Both the polysynaptic and monosynaptic components of the excitatory response of standard midbody RZ following cephalic nerve stimulation were inhibited in the presence of the kainate/quisqualate antagonist DNQX. These data suggest a role for glutamate as a transmitter in the neural circuitry from receptors of the leech prostomial lip to serotonergic RZ.

Distribution and partial characterization of CREB-like immunoreactivity in the medicinal leech Hirudo.

The presence and distribution of immunoreactivity to the cyclic AMP response element binding protein (CREB) were determined in the central nervous system (CNS) and in peripheral tissues of the medicinal leech Hirudo. Western blots revealed several CREB-immunoreactive (CREB-IR) bands including one whose molecular weight (43-44 kDa) was similar to mammalian CREB. The 43-44 kDa CREB-like protein was detected in nuclear extracts of the ventral nerve cord and was not observed following preincubation of the primary antiserum with the epitope sequence. CREB-like immunoreactivity was detected in extracts from each of six regions of the leech CNS, and in extracts from leech body wall musculature, crop, intestine, jaw musculature, pharynx, and salivary tissues. Whole mounts of leech ganglia revealed specific CREB-IR in a restricted population of neurons distributed throughout the leech CNS. Apparent homologues to a pair of CREB-IR dorsolateral neurons were observed in most ganglia along the ventral nerve cord. Several CREB-IR neurons exhibited segmental specificity. A number of neurons stained with an antiserum to the cyclic AMP response element modulator (CREM). These neurons showed no overlap in location with CREB-IR neurons, and this staining was not eliminated with a preabsorption control. Possible roles for a CREB-like protein in the leech are discussed.

Inhibitory actions of dopamine on Limulus visceral muscle involve a cyclic AMP-dependent mechanism.

1. The catecholamines dopamine, epinephrine and norepinephrine were detected in alumina extracts of Limulus midgut tissue using high performance liquid chromatography with electrochemical detection. Moderate levels of norepinephrine (28.2 +/- 2.1 ng/g) and dopamine (24.0 +/- 5.2 ng/g) were detected in the midgut, while epinephrine levels (7.4 +/- 0.9 ng/g) were less. Catecholamines were present in all regions along the longitudinal axis of the midgut, and norepinephrine and dopamine levels were highest in posterior regions. 2. Catecholamines decreased muscle tonus and inhibited spontaneous contractions of the Limulus midgut. Dopamine typically decreased spontaneous midgut activity at doses of 10(-8) M or greater, and produced inhibitory actions on all regions of the Limulus midgut. In some preparations epinephrine and norepinephrine elicited a secondary rhythmicity. The actions of dopamine opposed the excitatory effects produced by either proctolin or octopamine. 3. Catecholamines significantly elevated levels of cyclic AMP in Limulus midgut muscle rings. Dopamine (10(-5) M) increased cyclic AMP with a time course consistent with its physiological effects. Forskolin and several methyl xanthines increased Limulus midgut cyclic AMP levels and mimicked the inhibitory effects of dopamine on the isolated midgut preparation. Cyclic nucleotide analogues also produced dopamine-like effects on the isolated midgut preparation. Inhibition of cyclic nucleotide phosphodiesterase prior to addition of dopamine enhanced the effect of this amine to decrease baseline muscle tension. 4. The inhibitory effects of 10(-5) M dopamine on the midgut persisted in solutions of zero sodium and in the presence of tetrodotoxin. Zero calcium solutions gradually reduced spontaneous midgut activity and the effects of dopamine. Calcium channel blockers did not prohibit dopamine-induced relaxation.(ABSTRACT TRUNCATED AT 250 WORDS)

Mechanism for amine modulation of the neurogenic Limulus heart: evidence for involvement of cAMP.

The role of cyclic nucleotides as intracellular second messengers mediating the excitatory chronotropic and inotropic actions of octopamine (OCT) and dopamine (DA) on the neurogenic Limulus heart was investigated. Tissue levels of cAMP, but not cGMP, were significantly increased in isolated cardiac ganglia and cardiac muscle following 10 min exposure to 10(-5) M OCT or 10(-5) M DA. In both tissues, OCT elicited larger increases in cAMP than did DA. Amine-induced cAMP accumulation in the cardiac ganglion and in the cardiac muscle was prevented by the alpha-adrenergic blocker phentolamine. The adenylate cyclase activator forskolin and the phosphodiesterase inhibitor IBMX produced amine-like chronotropic and inotropic effects when applied to the isolated heart preparation. However, the kinetics of the responses differed for the two agents. Additional pharmacological agents (RO-20-1724, papaverine, SQ 20,009, and 8-parachloro-phenylthio cAMP) also had amine-like effects but to a lesser extent. The chronotropic, but not inotropic, effects of OCT and DA were potentiated in the presence of IBMX. These data suggest that a cAMP-dependent mechanism underlies the excitatory effects of the neuromodulators OCT and DA on the Limulus heart.

Second-messenger systems underlying amine and peptide actions on cardiac muscle in the horseshoe crab Limulus polyphemus.

The biochemical mechanisms by which octopamine, catecholamines and the peptide proctolin exert their actions on Limulus cardiac muscle were investigated. Amines produced long-lasting increases in the amplitude of contractions evoked by electrical stimulation. At 10(-5) mol l-1, the apparent order of potency for amine-induced increases in evoked contraction amplitude was dopamine approximately equal to octopamine greater than norepinephrine approximately equal to epinephrine. At this dose, amines produced long-lasting increases in the levels of cyclic AMP (octopamine greater than dopamine approximately equal to norepinephrine approximately equal to epinephrine), but not of cyclic GMP, in Limulus cardiac muscle. Like the amines, the adenylate cyclase activator forskolin enhanced cardiac muscle contractility and increased levels of cyclic AMP, but not of cyclic GMP. The phosphodiesterase inhibitor IBMX produced a transient increase in cardiac muscle contractility, but typically produced long-lasting negative inotropy. This agent increased levels of both cyclic AMP and cyclic GMP in Limulus cardiac muscle. Proctolin and the protein kinase C activator phorbol dB increased the contraction amplitude of the intact heart and the electrically stimulated myocardium. These compounds, as well as dopamine, elicited sustained contractures and rhythmic contractions when applied to deganglionated Limulus cardiac muscle rings. Unlike the amines, proctolin and phorbol dB did not increase cardiac muscle cyclic AMP levels. These results suggest that several second-messenger systems may be utilized by amines and peptides to produce excitatory actions on cardiac muscle fibers of the Limulus heart. Cyclic AMP appears to be an important second messenger underlying the effects of amines to enhance cardiac muscle contractility. Pharmacological data suggest that proctolin may alter cardiac muscle contractility and excitability by a mechanism which involves the phosphatidylinositol pathway. Dopamine, unlike the other amines, produces a number of proctolin-like effects and may activate both the cyclic AMP and the phosphatidylinositol systems in Limulus cardiac muscle.

Involvement of cyclic AMP in multiple, excitatory actions of biogenic amines on the cardiac ganglion of the horseshoe crab Limulus polyphemus.

Cyclic AMP appears to be involved in several excitatory actions of amines on neurones of the Limulus cardiac ganglion. Amines selectively increase levels of cardiac ganglion cyclic AMP with a magnitude and time course similar to that observed for amine-induced excitation of cardiac ganglion burst rate. With respect to either the physiological or biochemical effect, the apparent order of potency is octopamine greater than epinephrine approximately dopamine greater than norepinephrine. Elevation of cardiac ganglion cyclic AMP levels by octopamine or dopamine is dose-dependent and is potentiated by the phosphodiesterase inhibitor 3-isobutyl 1-methylxanthine (IBMX). Several pharmacological agents which influence cyclic nucleotide metabolism, including forskolin, IBMX and 8-substituted cyclic AMP analogues, have amine-like effects on the Limulus cardiac ganglion. These effects include increased burst rate of the isolated cardiac ganglion and decreased burst duration, interburst interval and number of spikes per burst in follower neurones. Forskolin and IBMX increase levels of cardiac ganglion cyclic AMP, and IBMX also increases cyclic GMP levels in this tissue. Amines, forskolin and IBMX have direct effects on follower neurones pharmacologically isolated from pacemaker cell input. Octopamine, forskolin and IBMX depolarize follower neurones, while dopamine hyperpolarizes these cells. Amines, forskolin and IBMX elicit burst-like potentials in follower neurones, and increase the size of evoked, unitary junction potentials recorded in cardiac muscle fibres. These pharmacological and biochemical data suggest that multiple, excitatory effects of biogenic amines on the Limulus cardiac ganglion are mediated by simultaneous increases in cyclic AMP at several loci within this neural network.

Comparison of silicone and latex catheters in the development of urethral stricture after cardiac surgery.

In an earlier series of 100 men followed up for between 3 and 12 months after coronary artery bypass grafting, the incidence of urethral stricture was 2%. These patients have now been followed up for between 15 and 24 months and the incidence of urethral stricture has risen to 5.2%. Latex catheters were used in these patients. A separate group of 117 men underwent coronary artery bypass grafting and were catheterised with silicone catheters. They were followed up for between 12 and 28 months and no urethral strictures were found. It is recommended that silicone catheters be used routinely for short-term catheterisation in men undergoing cardiac bypass surgery.

Urethral stricture following aortic graft surgery.

The incidence of urethral stricture following aortic graft surgery was investigated retrospectively in a group of 127 men. They had been catheterised prior to their operation with latex catheters. One man had been treated for a stricture 9 months after his operation. Information about post-operative urinary symptoms was obtained from a further 94 surviving patients by postal questionnaires sent at least 12 months following surgery. Eight patients underwent urological review including, where indicated, urethrography under antibiotic cover; 3 patients were identified with urethral strictures, an incidence of 3.2%. This is similar to that occurring after cardiac surgery, and urethral strictures should be considered a hazard whenever catheters are used in surgical procedures.

Acute aortocaval fistula in association with ruptured aortic aneurysm.

Eight cases of aortocaval fistula are described. In all of them operation was performed with repair of the fistula and replacement of the aneurysm by a prosthetic graft. In seven cases the cause was rupture of an atherosclerotic aneurysm but one case followed rupture of a re-entrant dissecting aortic aneurysm. A fistula is most easily recognized by the sensation of a palpable thrill over the aorta during operation together with systemic venous congestion and a high central venous pressure before aortic clamps are applied. Diagnosis of the fistula before surgery or its recognition during operation and before opening the aorta usually leads to a successful outcome.

The delay in recovery from fast inactivation in skeletal muscle sodium channels is deactivation.

1. Using macropatch techniques, we tested the assumption that deactivation underlies the observed delay in the onset to recovery from fast inactivation by comparing open-state deactivation to recovery delay for rat skeletal muscle mutations R1441C and R1441P. 2. Deactivation kinetics from the open state were determined from the exponential decay of tail currents. R1441C and R1441P prolonged open-state deactivation, with the greatest effect produced by R1441P. 3. Delays in the onset to recovery from fast inactivation for R1441P and for R1441C were abbreviated compared to those for rSkM1. Recovery delay was longer in R1441P than R1441C at voltages more negative than -110 mV. Recovery from inactivation exhibited a voltage dependence which, unlike delay, saturated at depolarized voltages. Recovery rate constants were increased to a similar extent for R1441C and R1441P at -150 to -120 mV compared to rSkM1. 4. These results indicate that the delay in the onset to recovery from fast inactivation in skeletal muscle sodium channels is due to deactivation. Lessening of charge immobilization for R1441C and R1441P may contribute to observed biophysical defects underlying the hyperexcitability of muscle fibers containing paramyotonia congenita mutations. The second stage of recovery from fast inactivation may be affected differentially by these mutations.