Adrenomedullin influences dissociated rat area postrema neurons

The area postrema (AP) is one of a specialized group of central nervous system (CNS) structures devoid of a significant blood-brain barrier (BBB), collectively known as the circumventricular organs (CVO). While peptides are normally excluded from access to most regions of the CNS, the AP contains neurons with a high density of receptors for many circulating peptides, very likely including those for adrenomedullin (AM). In this study, whole-cell patch-clamp recordings were obtained from 114 dissociated rat AP neurons. The mean resting membrane potential (RMP) of these neurons (n=79) was -54.3+/-0.8 mV, the mean input resistance (IR) was 3.1+/-0.2 GOmega and the spike amplitude of neurons included in this study was always greater than 90 mV. Current-clamp studies showed that bath application of AM depolarized 39.2% (31 of 79) and hyperpolarized 45.6% (36 of 79) of neurons tested. Both effects were found to be concentration dependent from 10(-12) to 10(-7) M. These data support the idea that specific populations of CNS neurons within the AP are directly influenced by AM and support the concept that AM may act at AP to influence central autonomic control. We also examined the roles of specific ion channels in regulating the AM-induced excitability of AP neurons through voltage-clamp studies. These experiments suggest potential actions of AM in modulating voltage gated calcium channels, effects which have the additional consequence of inhibiting calcium activated potassium conductances (I(K(Ca))). These data demonstrate direct effects of AM on dissociated AP neurons and identify ion channels, the modulation of which, may underlie these effects.     

Participation of the area postrema in cardiovascular control in the dog

Investigations have demonstrated that the pressor effects of low-dose intravertebral angiotensin II (Ang II) in the dog are mediated by the area postrema (AP). Chronic ablation of the AP has been shown to produce both mild hypotension and blunting of the pressor effects of peripherally administered Ang II, which suggests a tonic influence by this structure on the regulation of blood pressure. These findings motivated a correlated series of neurophysiological and anatomical studies to characterize further the AP pressor pathway. The pressor response to electrical stimulation of the AP was shown to be mediated by increased central sympathetic outflow, as shown previously for the response to intravertebral Ang II, and unopposed by the central nervous system baroreflex pathways. Neuroanatomical investigations demonstrated a three-layer structure in the dog's AP, with efferent projections into the medial nucleus tractus solitarii bilaterally. These studies have provided new evidence about the functional and structural mechanisms by which the AP participates in cardiovascular regulation.     

Effects of Extracellular Diadenosine Tetraphosphate on Action Potentials in the Atrial and Ventricular Myocardium of the Rat Heart during Early Postnatal Ontogenesis

Diadenosine tetraphosphate (AP4A) belongs to a wide group of naturally derived endogenous purine compounds that have recently been considered as new neurotransmitters in the autonomic nervous system. It has been shown that AP4A induces inhibitory effects and modulates adrenergic control in the heart of adult mammals. Nevertheless, the physiological significance of AP4A in early postnatal development, when sympathetic innervation remains yet immature, has not been investigated. The aim of the present study was to elucidate the effects of AP4A on the heart bioelectrical activity in early postnatal ontogenesis. Action potentials (AP) were recorded using the standard microelectrode technique in multicellular isolated right atrial (RA), left atrial (LA), and ventricle (RV) preparations from male Wistar rats at postnatal days 1, 14, and 21 and from 60-day animals that were considered as adults. The application of AP4A caused significant reduction of AP duration in atrial (RA and LA) preparations from rats of all ages. Also, AP4A caused significant AP shortening in RV preparations from rats of various ages; however, the effect was more pronounced in 21-day-old and adult rats. AP4A failed to alter automaticity of RA preparations from the rats at postnatal days 1, 14, and 21 and weakly decreased spontaneous rhythm in RA preparations from the adult rats. The effect of AP4A was partially abolished by P2-receptor blocker PPADS in LA preparations from both 21-dayold and adult rats, while it failed to suppress AP4A-caused AP shortening in preparations from 1- and 14-dayold animals. Thus, extracellular AP4A causes shortening of AP both in the atrial and ventricular myocardium in the rats of early postnatal ontogenesis and in adults. The effect of AP4A depends on age only for ventricular myocardium where it may be attributed with growing contribution of diadenosine polyphosphates to the control of myocardium inotropy.     

The area postrema: a cardiovascular control centre at the blood-brain interface?

The area postrema (AP) is one of the circumventricular organs of the brain and as such it is highly vascular and lacks the normal blood-brain barrier. Anatomical tracing studies have demonstrated afferent projections to AP originating from the paraventricular nucleus, lateral parabrachial nucleus (l-PBN), nucleus tractus solitarius (NTS), as well as the vagus nerve. AP neurons have been shown to project primarily to l-PBN, and NTS. Receptor localization studies have reported dense aggregations of many specific peptide receptors in AP including those for angiotensin II (ANG), atrial natriuretic peptide (ANP), and endothelin (ET). Electrical stimulation studies have shown that activation of AP neurons at low frequencies (less than 15 Hz) results in decreases in blood pressure and heart rate, while higher frequency (greater than 20 Hz) stimulation causes increases in blood pressure. These low frequency effects on blood pressure and heart rate appear to result from activation of separate components of the autonomic nervous system. Extracellular single unit recordings have identified two functionally separate populations of AP neurons: one responsive to circulating ANG and a second apparently responsive to changes in blood pressure. In addition, AP neurons are activated by increases in circulating ET. Afferent inputs to AP neurons from 1-PBN have separate excitatory (12% of AP neurons) or inhibitory (12% of AP neurons) effects on a relatively small proportion of AP neurons. In contrast, preliminary evidence suggests a much more broadly distributed excitatory input to approximately 70% of tested AP neurons originating from the aortic depressor nerve. These studies provide considerable evidence implicating the AP as a significant neural structure regulating the cardiovascular system.     

Dissociation of Na+,K+-ATPase inhibition from digitalis inotropy.

Recent findings from our laboratory as well as those of other laboratories do not support the postulation that the mechanism of the positive inotropic action of digitalis is due to inhibition of NA,K-ATPase. Using short-acting digitalis steroids and drug washout experiments, in isolated myocardial preparations, it has been demonstrated that Na,K-ATPase isolated from such preparations is still significantly inhibited, whereas the positive inotropic effect is no longer present. Also, based on kinetic measurements the two exponential rate constants observed for drug half-life, a rapid and slow phase, were found to be associated, respectively, with the very short inotropic half-life and the very long enzyme inhibition half-life. In addition, a dissociation of the transient inotropic effects of digitalis was observed from the long lasting cardiotoxic effects of digitalis during drug washout. Moreover, a temporal correlation was noted between the persistent inhibitory effects of digitalis on Na,K-ATPase and the persistent cardiotoxic effects of digitalis. Therefore, it is concluded that inhibition of Na,K-ATPase is not responsible for the positive inotropic action of digitalis, but may be the mechanism, at least in part, for certain cardiotoxic effects of digitalis.     

Antazoline therapy of recurrent refractory supraventricular arrhythmias--a preliminary report.

Seven patients with chronic or recurrent supraventricular tachyarrhythmias were selected for a trial of antazoline therapy because sinus rhythm or a controlled ventricular response could not be achieved with quinidine, procainamide, digitalis or propranolol. Sinus rhythm was established by either intravenous administration of antazoline or direct-current countershock, and has been maintained in all for 4 to 16 months by oral administration of antazoline. Side effects were minor. Antazoline is a sufficiently promising antiarrhythmic agent to warrant large-scale controlled studies.     

GAP-43 is expressed by nonmyelin-forming Schwann cells of the peripheral nervous system.

Recently it has been demonstrated that the growth-associated protein GAP-43 is not confined to neurons but is also expressed by certain central nervous system glial cells in tissue culture and in vivo. This study has extended these observations to the major class of glial cells in the peripheral nervous system, Schwann cells. Using immunohistochemical techniques, we show that GAP-43 immunoreactivity is present in Schwann cell precursors and in mature non-myelin-forming Schwann cells both in vitro and in vivo. This immunoreactivity is shown by Western blotting to be a membrane-associated protein that comigrates with purified central nervous system GAP-43. Furthermore, metabolic labeling experiments demonstrate definitively that Schwann cells in culture can synthesize GAP-43. Mature myelin-forming Schwann cells do not express GAP-43 but when Schwann cells are removed from axonal contact in vivo by nerve transection GAP-43 expression is upregulated in nearly all Schwann cells of the distal stump by 4 wk after denervation. In contrast, in cultured Schwann cells GAP-43 is not rapidly upregulated in cells that have been making myelin in vivo. Thus the regulation of GAP-43 appears to be complex and different from that of other proteins associated with nonmyelin-forming Schwann cells such as N-CAM, glial fibrillary acidic protein, A5E3, and nerve growth factor receptor, which are rapidly upregulated in myelin-forming cells after loss of axonal contact. These observations suggest that GAP-43 may play a more general role in the nervous system than previously supposed.     

Effects of exogenous fatty acids and cholesterol on aminopeptidase activities in rat astroglia

Several studies have addressed the interaction between fatty acids and lipids with central nervous system peptides. Because aminopeptidases (AP) are involved in the regulation of neuropeptides, this work studies several AP expressed in cultured astroglia, after exogenous addition of oleic and linoleic fatty acids and cholesterol to the culture medium. Alanyl-AP, arginyl-AP, cystyl-AP, leucyl-AP, tyrosyl-AP and pyroglutamyl-AP activities were analysed in whole cells using the corresponding aminoacyl-beta-naphthylamides as substrates. Oleic acid inhibits alanyl-AP, cystyl-AP and leucyl-AP activities, whereas linoleic acid inhibits alanyl-AP, arginyl-AP and tyrosyl-AP activities. Neither oleic acid nor linoleic acid modifies pyroglutamyl-AP activity. In contrast, cholesterol increases arginyl-AP, cystyl-AP, leucyl-AP, tyrosyl-AP and pyroglutamyl-AP activities, although it does not modify alanyl-AP activity. The changes reported here suggest that oleic and linoleic fatty acids and cholesterol can modulate peptide activities via their degradation route involving aminopeptidases; each of them being differentially regulated.     

Transplantation of specific human astrocytes promotes functional recovery after spinal cord injury

Repairing trauma to the central nervous system by replacement of glial support cells is an increasingly attractive therapeutic strategy. We have focused on the less-studied replacement of astrocytes, the major support cell in the central nervous system, by generating astrocytes from embryonic human glial precursor cells using two different astrocyte differentiation inducing factors. The resulting astrocytes differed in expression of multiple proteins thought to either promote or inhibit central nervous system homeostasis and regeneration. When transplanted into acute transection injuries of the adult rat spinal cord, astrocytes generated by exposing human glial precursor cells to bone morphogenetic protein promoted significant recovery of volitional foot placement, axonal growth and notably robust increases in neuronal survival in multiple spinal cord laminae. In marked contrast, human glial precursor cells and astrocytes generated from these cells by exposure to ciliary neurotrophic factor both failed to promote significant behavioral recovery or similarly robust neuronal survival and support of axon growth at sites of injury. Our studies thus demonstrate functional differences between human astrocyte populations and suggest that pre-differentiation of precursor cells into a specific astrocyte subtype is required to optimize astrocyte replacement therapies. To our knowledge, this study is the first to show functional differences in ability to promote repair of the injured adult central nervous system between two distinct subtypes of human astrocytes derived from a common fetal glial precursor population. These findings are consistent with our previous studies of transplanting specific subtypes of rodent glial precursor derived astrocytes into sites of spinal cord injury, and indicate a remarkable conservation from rat to human of functional differences between astrocyte subtypes. In addition, our studies provide a specific population of human astrocytes that appears to be particularly suitable for further development towards clinical application in treating the traumatically injured or diseased human central nervous system.     

Vertebrate anteroposterior patterning: the Xenopus neurectoderm as a paradigm

This review discusses formation of the vertebrate anteroposterior (AP) axis, focusing on the dorsal ectoderm, which gives rise to the nervous system, using the frog Xenopus as a model. After summarizing classical models of AP neural patterning, we describe recent molecular studies that are encouraging re-examination of these models. Such studies have shown that AP ectodermal patterning occurs by the onset of gastrulation, much earlier than previously thought. The identity of tissues that determine AP pattern is discussed, and the definition of the Organizer is reconsidered. The activity of factors secreted by inducing tissues in early patterning decisions is assessed and formulated into a revised model for Xenopus AP neural patterning. Finally, AP ectodermal patterning in Xenopus dorsal ectoderm is compared to that of other germ layers, and to other vertebrates.     

白介素1β抑制培养的大鼠皮层神经元钠电流峰值和动作电位幅度

白介素1β(interleukin-1β,IL-1β)是重要的促炎细胞因子,在中枢神经系统的生理学和病理学过程中发挥关键作用.电压门控钠通道是可兴奋细胞电学活动的基础,控制神经元的兴奋性和动作电位.最近的研究又显示了IL-1β与电压门控通道之间的相互作用.为考察中枢神经元中IL-1β与电压门控钠通道之间的相互作用,本研...     

Association of the positive inotropic action of ouabain with a second species of digitalis receptors

Studies were conducted to determine whether Na-K ATPase or a second species of digitalis receptors in canine cardiac sarcolemma membrane preparations is associated with the positive inotropic action of nontoxic concentrations of ouabain. [3H]ouabain association and dissociation experiments using highly enriched sarcolemma preparations from canine ventricle indicate the presence of two species of ouabain binding receptors. Ouabain binding to Na-K ATPase of the sarcolemma preparation requires supporting ligands and is characterized by fast association and very slow dissociation in vitro. The second species of digitalis receptor does not require supporting ligands for ouabain binding and is characterized by slow association and fast dissociation. To determine which species of digitalis receptor is associated with the positive inotropic action of digitalis, ouabain washout experiments were conducted using various isolated canine myocardial preparations. Washout of the positive inotropic effects of 1.2-2.4 X 10(-7) M ouabain gave half-life values of 1.5-2.0 h for the various myocardial preparations. [3H]ouabain dissociation from the second species of digitalis receptors gave half-life values of 1.7-1.8 h, whereas dissociation from the sarcolemma Na-K ATPase gave half-life values of 8.9-9.3 h for the various sarcolemma preparations utilized. Therefore, based on similarities in half-life values between ouabain inotropy and [3H]ouabain dissociation from the second class of digitalis receptors, it is postulated that the positive inotropic action of digitalis glycosides is associated with the second species of digitalis receptors in the sarcolemma and not with the digitalis inhibitory receptor of Na-K ATPase for nontoxic concentrations of digitalis.     

Epidermal growth factor promotes oligodendrocyte process formation and regrowth after injury

Oligodendrocytes (OLs) form myelin within the central nervous system and are targets in numerous demyelinating diseases and injuries. OLs grown in culture maintain the developmental timetable which occurs in vivo and mature into cells with a relatively normal phenotype. In this study, cultured cells are used to test whether EGF can modulate process formation in OLs both before and after transection injury. EGF had no effect on the formation of new processes by OLs at any stage of development. To test the effect of EGF on process outgrowth after injury, mature OLs were selected and injured by laser transection of a single process, then imaged at 24-h intervals for 120 h. EGF promoted the recovery and regrowth of injured processes and also significantly increased outgrowth in uninjured processes. As well, it increased the number of new sprouts formed by OLs after injury. Results suggest that the effects of EGF on process outgrowth are a consequence of EGF interaction with a signaling pathway that is specifically activated within injured OLs. The potent effect of EGF on OL process formation after an injury suggests that modulation of the signaling pathways involved might provide a mechanism to promote remyelination.     

Activity of the alternative complement pathway during mycobacterial infections in inbred mice

The relationship between kinetic activity of the alternative pathway (AP) of complement and susceptibility to Mycobacterium bovis (BCG) and M. fortuitum was examined in inbred mice. After subcutaneous injection of BCG, the organisms were mostly contained by the draining lymph nodes, with minimal effects on spleen and no apparent relationship with serum AP. After intravenous injection of BCG or M. fortuitum, male mice, which had a more effective AP than female mice, showed lower spleen bacterial counts. AP kinetics became faster in mice with high spleen bacterial counts and slower in mice with low counts, suggesting that infection or inflammatory processes affected AP. These experiments suggest that if tuberculosis is confined to tissues and draining lymph nodes AP plays no part in pathogenesis or host resistance, but AP might reduce the infectivity of low numbers of organisms spreading by blood or lymph from a primary focus of infection.     

Forced Desynchrony Reveals Independent Contributions of Suprachiasmatic Oscillators to the Daily Plasma Corticosterone Rhythm in Male Rats

The suprachiasmatic nucleus (SCN) is required for the daily rhythm of plasma glucocorticoids; however, the independent contributions from oscillators within the different subregions of the SCN to the glucocorticoid rhythm remain unclear. Here, we use genetically and neurologically intact, forced desynchronized rats to test the hypothesis that the daily rhythm of the glucocorticoid, corticosterone, is regulated by both light responsive and light-dissociated circadian oscillators in the ventrolateral (vl-) and dorsomedial (dm-) SCN, respectively. We show that when the vlSCN and dmSCN are in maximum phase misalignment, the peak of the plasma corticosterone rhythm is shifted and the amplitude reduced; whereas, the peak of the plasma adrenocorticotropic hormone (ACTH) rhythm is also reduced, the phase is dissociated from that of the corticosterone rhythm. These data support previous studies suggesting an ACTH-independent pathway contributes to the corticosterone rhythm. To determine if either SCN subregion independently regulates corticosterone through the sympathetic nervous system, we compared unilateral adrenalectomized, desynchronized rats that had undergone either transection of the thoracic splanchnic nerve or sham transection to the remaining adrenal. Splanchnicectomy reduced and phase advanced the peak of both the corticosterone and ACTH rhythms. These data suggest that both the vlSCN and dmSCN contribute to the corticosterone rhythm by both reducing plasma ACTH and differentially regulating plasma corticosterone through an ACTH- and sympathetic nervous system-independent pathway.     

Effect of Propranolol on Exercise Tolerance of Patients with Atrial Fibrillation

Six patients with atrial fibrillation who were taking digitalis were exercised before and after 30 mg. of propranolol twice daily. Though there was a lower pulse rate at rest and on exercise in all patients, three suffered deterioration of exercise tolerance. It is concluded that propranolol does not improve the exercise tolerance of patients with atrial fibrillation whose resting ventricular rate is controlled with digitalis.     

Intracellular Ca2+ release underlies the development of phase 2 in mouse ventricular action potentials

The ventricular action potential (AP) is characterized by a fast depolarizing phase followed by a repolarization that displays a second upstroke known as phase 2. This phase is generally not present in mouse ventricular myocytes. Thus we performed colocalized electrophysiological and optical recordings of APs in Langendorff-perfused mouse hearts founding a noticeable phase 2. Ryanodine as well as nifedipine reduced phase 2. Our hypothesis is that a depolarizing current activated by Ca(2+) released from the sarcoplasmic reticulum (SR) rather than the "electrogenicity" of the L-type Ca(2+) current is crucial in the generation of mouse ventricular phase 2. When Na(+) was partially replaced by Li(+) in the extracellular perfusate or the organ was cooled down, phase 2 was reduced. These results suggest that the Na(+)/Ca(2+) exchanger functioning in the forward mode is driving the depolarizing current that defines phase 2. Phase 2 appears to be an intrinsic characteristic of single isolated myocytes and not an emergent property of the tissue. As in whole heart experiments, ventricular myocytes impaled with microelectrodes displayed a large phase 2 that significantly increases when temperature was raised from 22 to 37°C. We conclude that mouse ventricular APs display a phase 2; however, changes in Ca(2+) dynamics and thermodynamic parameters also diminish phase 2, mostly by impairing the Na(+)/Ca(2+) exchanger. In summary, these results provide important insights about the role of Ca(2+) release in AP ventricular repolarization under physiological and pathological conditions.     

Digitoxigenin, a digitalis steroid, induces meiotic maturation of Xenopus laevis oocytes

Digitoxigenin, a C23 digitalis steroid induces meiotic maturation of Xenopus oocyte. The dose of digitoxigenin which induces half maximal response is 3.3 +/- 2.10(-5)M. In contrast the conjugated digitalis steroid, digitoxin (digitoxigenine + 3 digitoxoses) never triggered maturation at any of the doses tested. These experiments which show that only free digitoxigenin mimics progesterone action, suggest that both digitoxigenin and progesterone possess a common initial site of action which is not localized at the level of the outer leaflet of the oocyte plasma membrane.     

Rapid corticosterone-induced impairment of amplectic clasping occurs in the spinal cord of roughskin newts (taricha granulosa)

Courtship clasping, a reproductive behavior in male roughskin newts (Taricha granulosa), is rapidly blocked by an action of corticosterone (CORT) at a specific neuronal membrane receptor. The CORT-induced impairment of clasping in behaving newts appears to be mediated partly by an elimination of clasping-related activity in medullary reticulospinal neurons. Previous studies of rapid CORT actions in Taricha have focused on the brain, so existence of CORT action in the spinal cord or peripheral nervous system has not been assessed. The present study used newts with a high cervical spinal transection to examine potential spinal or peripheral CORT effects on clasping by the hindlimbs in response to pressure on the cloaca. Spinal transection causes clasps elicited by cloacal stimulation to be very sustained beyond the termination of the eliciting stimulus. In spinally transected newts, CORT caused a dose-dependent depression in the duration as well as quality of the clasp that appeared within 10 min of injection. CORT selectively impaired the usual sustained maintenance of a clasp after termination of cloacal stimulation, but not clasp elicitation during stimulation. These effects were not produced by dexamethasone, a synthetic glucocorticoid that binds poorly to the CORT membrane receptor. The CORT effect on clasp maintenance but not clasp elicitation implies selective action on an intraspinal generator for clasping but not on sensory or efferent neuromuscular aspects of the response. These results indicate the presence in the newt spinal cord of the CORT membrane receptor that exerts functional effects distinctly different from those on the brainstem.