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1.
Voltage-dependent calcium and calcium-activated potassium currents of a molluscan photoreceptor 总被引:5,自引:1,他引:4
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Two-microelectrode voltage clamp studies were performed on the somata of Hermissenda Type B photoreceptors that had been isolated by axotomy from all synaptic interaction as well as any impulse-generating (i.e., active) membrane. In the presence of 2-10 mM 4-aminopyridine (4-AP) and 100 mM tetraethylammonium ion (TEA), which eliminated two previously described voltage-dependent potassium currents (IA and the delayed rectifier), a voltage-dependent outward current was apparent in the steady state responses to command voltage steps more positive than -40 mV (absolute). This current increased with increasing external Ca++. The magnitude of the outward current decreased and an inward current became apparent following EGTA injection. Substitution of external Ba++ for Ca++ also made the inward current more apparent. This inward current, which was almost eliminated after being exposed for approximately 5 min to a solution in which external Ca++ was replaced with Cd++, was maximally activated at approximately 0 mV. Elevation of external potassium allowed the calcium (ICa++) and calcium-dependent K+ (IC) currents to be substantially separated. Command pulses to 0 mV elicited maximal ICa++ but no IC because no K+ currents flowed at their new reversal potential (0 mV) in 300 mM K+. At a holding potential of -60 mV, which was now more negative than the potassium equilibrium potential, EK+, in 300 mM K+, IC appeared as an inward tail current after positive command steps. The voltage dependence of ICa++ was demonstrated with positive steps in 100 mM Ba++, 4-AP, and TEA. Other data indicated that in 10 mM Ca++, IC underwent pronounced and prolonged inactivation whereas ICa++ did not. When the photoreceptor was stimulated with a light step (with the membrane potential held at -60 mV), there was also a prolonged inactivation of IC. In elevated external Ca++, ICa++ also showed similar inactivation. These data suggest that IC may undergo prolonged inactivation due to a direct effect of elevated intracellular Ca++, as was previously shown for a voltage-dependent potassium current, IA. These results are discussed in relation to the production of training-induced changes of membrane currents on retention days of associative learning. 相似文献
2.
D L Alkon M Kubota J T Neary S Naito D Coulter H Rasmussen 《Biochemical and biophysical research communications》1986,134(3):1245-1253
Voltage-dependent K+ currents, IA and ICa2+-K+, across the soma membrane of the Hermissenda Type B photoreceptor, have been shown to remain reduced during retention of classically conditioned behavior. IA and ICa2+-K+ undergo prolonged reduction due to [Ca2+]i elevation produced by a single pairing of a light step with a command depolarization or by iontophoretic injection of Ca2+. One pathway which could contribute to the conversion of transient Ca2+-mediated reduction of K+ currents to the persistent reduction observed with conditioning is that involving C-kinase. To examine the role of C-kinase in the long-term regulation of K+ currents, isolated Type B somata were exposed to at least 25-30 minutes' incubation in artificial sea water (ASW) containing the C-kinase activators 1-oleoyl-2-acetyl-glycerol (OAG) or 12-deoxyphorbol 13-isobutyrate 20-acetate (DPBA) or control substances [e.g., distearyolglycerol (DiSG)]. After exposure to activator (but not to control solutions) and voltage-clamp conditions which caused elevation of cytosolic Ca2+, reductions of IA and ICa2+-K+ were observed which did not reverse (up to 3 hr), even after the activator was removed. Without conditions which induced elevation of cytosolic calcium prolonged incubation with the C-kinase activators had no effect on the membrane currents. Similar exposure of homogenates of the Hermissenda nervous system to OAG and Ca2+ caused enhanced phosphorylation of specific proteins, indicating the presence of C-kinase in the Hermissenda nervous system. 相似文献
3.
In previous studies elevation of intracellular Ca2+ was shown to cause prolonged reduction of two voltage-dependent K+ currents (IA and ICa2+-K+) across the membrane of the isolated Hermissenda photoreceptor, the type B cell (Alkon et al., 1982b; Alkon and Sakakibara, 1985). Here we show that iontophoretic injection of inositol trisphosphate (IP3), but not inositol monophosphate, also caused prolonged reduction of IA and ICa2+-K+. IP3 injection also caused reduction of a light-induced K+ current (also ICa2+-K+) but did not affect the voltage-dependent Ca2+ current, ICa2+, or the light-induced inward current, INa+, of the type B cell. IP3 injection caused similar effects on the K+ currents of the other type of Hermissenda photoreceptor, the type A cell. INA+ of the type A cell, unlike that of the type B cell, was, however, markedly increased following IP3 injection. The differences of IP3 effects on the two types of photoreceptors may be related to differences in regulation of ionic currents by endogenous IP3 as reflected by clear differences (before injection) in the magnitude of IA, ICa2+-K+, and INa+ between the two cell types. 相似文献
4.
E N Yamoah 《Biophysical journal》1997,72(1):193-203
Apart from their primary function as balance sensors, Hermissenda hair cells are presynaptic neurons involved in the Ca(2+)-dependent neuronal plasticity in postsynaptic B photoreceptors that accompanies classical conditioning. With a view to beginning to understand presynaptic mechanisms of plasticity in the vestibulo-visual system, a locus for conditioning-induced neuronal plasticity, outward currents that may govern the excitability of hair cells were recorded by means of a whole-cell patch-clamp technique. Three K+ currents were characterized: a 4-aminopyridine-sensitive transient outward K+ current (IA), a tetraethyl ammonium-sensitive delayed rectifier K+ current (IK,V), and a Ca(2+)-activated K+ current (IK,Ca). IA activates and decays rapidly; the steady-state activation and inactivation curves of the current reveal a window current close to the apparent resting voltage of the hair cells, suggesting that the current is partially activated at rest. By modulating firing frequency and perhaps damping membrane oscillations, IA may regulate synaptic release at baseline. In contrast, IK,V and IK,Ca have slow onset and exhibit little or no inactivation. These two K+ currents may determine the duration of the repolarization phase of hair-cell action potentials and hence synaptic release via Ca2+ influx through voltage-gated Ca2+ channels. In addition, IK,Ca may be responsible for the afterhyperpolarization of hair cell membrane voltage following prolonged stimulation. 相似文献
5.
Regulation of Hermissenda K+ Channels by Cytoplasmic and Membrane-Associated C-Kinase 总被引:3,自引:0,他引:3
D. L. Alkon S. Naito M. Kubota C. Chen B. Bank J. Smallwood P. Gallant H. Rasmussen 《Journal of neurochemistry》1988,51(3):903-917
Pharmacologic activation of endogenous protein kinase C (PKC) together with elevation of the intracellular Ca2+ level was previously shown to cause reduction of two voltage-dependent K+ currents (IA and ICa2+-K+) across the soma membrane of the type B photoreceptor within the eye of the mollusc Hermissenda crassicornis. Similar effects were also found to persist for days after acquisition of a classically conditioned response. Also, the state of phosphorylation of a low-molecular-weight protein was changed only within the eyes of conditioned Hermissenda. To examine the role of PKC in causing K+ current changes as well as changes of phosphorylation during conditioning (and possibly other physiologic contexts), we studied here the effects of endogenous PKC activation and exogenous PKC injection on phosphorylation and K+ channel function. Several phosphoproteins (20, 25, 56, and 165 kilodaltons) showed differences in phosphorylation in response to PKC activators applied to intact nervous systems or to isolated eyes. Specific differences were observed for membrane and cytosolic fractions in response to both the phorbol ester 12-deoxyphorbol 13-isobutyrate 20-acetate (DPBA) or exogenous PKC in the presence of Ca2+ and phosphatidylserine/diacylglycerol. Type B cells pretreated with DPBA responded to PKC injection with a persistent reduction of K+ currents. In the absence of DPBA, PKC injection also caused K+ current reduction only following Ca2+ loading conditions. However, the direct effect of PKC injection in the absence of DPBA was only to increase ICa2+-K+. According to a proposed model, the amplitude of the K+ currents would depend on the steady-state balance of effects mediated by PKC within the cytoplasm and membrane-associated PKC. The model further specifies that the effects on K+ currents of cytoplasmic PKC require an intervening proteolytic step. Such a model predicts that increasing the concentration of cytoplasmic protease, e.g., with trypsin, will increase K+ currents, whereas blocking endogenous protease, e.g., with leupeptin, will decrease K+ currents. These effects should be opposed by preexposure of the cells to DPBA. Furthermore, prior injection of leupeptin should block or reverse the effects of subsequent injection of PKC into the type B cell. All of these predictions were confirmed by results reported here. Taken together, the results of this and previous studies suggest that PKC regulation of membrane excitability critically depends on its cellular locus. The implications of such function for long-term physiologic transformations are discussed. 相似文献
6.
Elevated intracellular Ca++ concentration reduces the amplitude of an early, voltage-dependent K+ current (IA) in the Type B photoreceptor of Hermissenda crassicornis. Internal Ca++ is increased by activating a voltage and light-dependent Ca++ current present in these cells or by direct iontophoresis of Ca++ ions. Substitution of Ba++ for Ca++ or elimination of Ca++ from the sea water bathing the cells abolishes the reduction in IA during paired light and depolarizing voltage steps. The delayed K+ current (IB) in these cells is also reduced during paired light and voltage steps, but this decrease of IB is not affected by removal of extracellular Ca++. IB (but not IA), apparently much less dependent on intracellular Ca++ levels, is reduced by light alone. Ca++ iontophoresis also abolishes the light-dependent Na+ current, which recovers with a time course of minutes. 相似文献
7.
Two-microelectrode voltage-clamp measurements were made to determine the kinetics and voltage dependence of ionic currents across the soma membrane of the Hermissenda type B photoreceptor. The voltage-dependent outward potassium currents, IA and ICa(2+)-K+, the inward voltage-dependent calcium current, ICa2+ and the light-induced current, IIgt, were then described with Hodgkin-Huxley-type equations. The fast-activating and inactivating potassium current, IA, was described by the equation; IA(t) = gA(max)(ma infinity[1-exp(-t/tau ma)])3 x (ha infinity [1-exp(-t/tau ha)] + exp(-t/tau ha)) (Vm-EK), where the parameters ma infinity, ha infinity, tau ma, and tau ha are functions of membrane potential, Vm, and ma infinity and ha infinity are steady-state activation and inactivation parameters. Similarly, the calcium-dependent outward potassium current, ICa(2+)-K+, was described by the equation, ICa(2+)-K+ (t) = gc(max)(mc infinity(VC)(1-exp[-t/tau mc (VC)]))pc (hc infinity(VC) [1-exp(-t/tau hc)] + exp(-t/tau hc(VC)])pc(VC-EK). In high external potassium, ICa(2+)-K+ could be measured in approximate isolation from other currents as a voltage-dependent inward tail current following a depolarizing command pulse from a holding potential of -60 mV. A voltage-dependent inward calcium current across the type B soma membrane, ICa2+, activated rapidly, showed little inactivation, and was described by the equation: ICa2+ = gCa(max) [1 + exp](-Vm-5)/7]-1 (Vm-ECa), where gCa(max) was 0.5 microS. The light-induced current with both fast and slow phases was described by: IIgt(t) = IIgt1 + IIgt2 + IIgt3, IIgti = gIgti [1-exp(- ton/tau mi)] exp(-ton/tau hi)(Vm-EIgti) (i = 1, 2).(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
8.
Intracellularly injected tumor promoter phorbol esters (PhEs) that activate protein kinase C (PKC) increased the excitability and altered the postsynaptic responses of neurons of the motor cortex of awake cats. PhEs increased the amplitude and duration of EPSPs and decreased the amplitude and durations of IPSPs. No consistent changes in resting membrane parameters that would account for these modifications were found. Corresponding changes in peak excitatory and inhibitory postsynaptic currents (EPSCs, IPSCs) were measured directly with the single electrode voltage clamp technique. The changes lasted for 50 min or longer. Quantitative analysis of EPSCs in response to ventrolateral thalamic stimulation and IPSCs in response to pyramidal tract stimulation made in a subgroup of fast PT cells suggested that PhE acted within the injected neuron rather than presynaptically to alter the synaptic currents. PhE also reduced a voltage-dependent, 3-aminopyridine sensitive fast outward current (IA) and an apamin and EGTA sensitive slow outward current (IK(Ca]. Control injections of a phorbol ester that did not activate PKC failed to induce changes in synaptic responses or resting membrane properties. These observations provide the first evidence that activation of PKC, in vivo, can induce long-lasting changes in synaptic responses of neocortical neurons by direct modification of postsynaptic ion channel conductivities. 相似文献
9.
A series of studies on Hermissenda classical conditioning has lead to a discovery that the biophysical events (accumulation of Ca2+ and depolarization in B cell) found during memory acquisition are clearly distinct from those (suppression of K-currents, IA and ICa2+K+) detected in the retention phase of memory. Biochemical analysis of eyes isolated shortly after (a few hours) training revealed increased phosphorylation of a 20,000 M.W. protein which is very likely one of the substrates for both Ca/CaM-dependent protein kinase and C-kinase and possibly a locus of convergence for conditioned stimulus and unconditioned stimulus pathways. Furthermore, conditioning-specific changes in the two K+ currents have been reproduced by simultaneous activation of the CaM-kinase pathway (via iontophoretic injection of CaM-kinase II plus Ca2+-load or IP3 injection) and the C-kinase pathway (via bath application of phorbol-ester or diacylglycerol analog plus Ca2+-load). Therefore, synergistic interaction between the two Ca2+-dependent phosphorylation systems in the identified B cell is considered to be critically important for acquisition of associative memory. Evidence also has been obtained for similar biophysical changes and molecular mechanisms during retention of classical conditioning in the mammalian brain. Further work will be needed to uncover the biochemical mechanism(s) responsible for transforming short-term into long-lasting memory. 相似文献
10.
Neutralizing monoclonal antibody against ras oncogene product p21 which impairs guanine nucleotide exchange. 总被引:13,自引:7,他引:6
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S Hattori D J Clanton T Satoh S Nakamura Y Kaziro M Kawakita T Y Shih 《Molecular and cellular biology》1987,7(5):1999-2002
The neutralizing monoclonal antibody Y13-259 severely hampers the nucleotide exchange reaction between p21-bound and exogenous guanine nucleotides but does not interfere with the association of GDP to p21. These results suggest that the nucleotide exchange reaction is critical for p21 function. Interestingly, the v-ras p21 has a much faster dissociation rate than the p21 of the c-ras proto-oncogene. 相似文献
11.
Manabu Sakakibara Carlos Collin Alan Kuzirian Daniel L. Alkon Eliahu Heldman Shigetaka Naito Izja Lederhendler 《Journal of neurochemistry》1987,48(2):405-416
Type B photoreceptors of the nudibranch mollusc Hermissenda crassicornis receive excitatory synaptic potentials (EPSPs) whose frequency is controlled by potential changes of a neighboring cell known as the S optic ganglion cell which is thought to be electrically coupled to the presynaptic source of these EPSPs, the E optic ganglion cell. The frequency of the EPSPs increases when a conditioned stimulus (light) is paired with an unconditioned stimulus (rotation) during acquisition of a Pavlovian conditioned response. The results of the present study are consistent with an adrenergic origin for these EPSPs. Noradrenergic agonists (greater than 100 microM), norepinephrine and clonidine, only slightly depolarize the type B cell but clearly prolong its depolarizing response to light. Serotonin, by contrast, causes hyperpolarization of the type B cell's resting potential as well as after a light step. Clonidine reduces voltage-dependent outward K+ currents (IA, an early current, ICa2+-K+, a late Ca2+-dependent current) that control the type B cell's excitability (and thus its light response and membrane potential). These effects of clonidine are reduced or blocked by the alpha 2-receptor antagonist, yohimbine (0.5 microM), but not the alpha 1-blocker, prazosin. The same yohimbine concentration also blocked depolarizing synaptic excitation of the type B cell in response to depolarization of a simultaneously impaled S optic ganglion cell. Histochemical techniques (both the glyoxylic acid method of de la Torre and Surgeon and the formaldehyde-induced fluorescence or Falck-Hillarp method) demonstrated the presence of a biogenic amine(s) within a single neuron in each optic ganglion as well as three or four cells within the vicinity of previously identified visual interneurons. No serotonergic neurons were found within the optic ganglion or in proximity to visual interneurons. A clonidine-like synaptic effect on type B cells, therefore, could amplify conditioning-specific changes of membrane currents by increasing type B depolarization and possibly, as well, by elevating intracellular second messengers. 相似文献
12.
Voltage-gated transient currents in bovine adrenal fasciculata cells. II. A-type K+ current 总被引:1,自引:0,他引:1
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《The Journal of general physiology》1993,102(2):239-255
In whole cell patch clamp recordings on enzymatically dissociated adrenal zona fasciculata (AZF) cells, a rapidly inactivating A-type K+ current was observed in each of more than 150 cells. Activation of IA was steeply voltage dependent and could be described by a Boltzmann function raised to an integer power of 4, with a midpoint of -28.3 mV. Using the "limiting logarithmic potential sensitivity," the single channel gating charge was estimated to be 7.2 e. Voltage-dependent inactivation could also be described by a Boltzmann function with a midpoint of -58.7 mV and a slope factor of 5.92 mV. Gating kinetics of IA included both voltage-dependent and -independent transitions in pathways between closed, open, and inactivated states. IA activated with voltage-dependent sigmoidal kinetics that could be fit with an n4h formalism. The activation time constant, tau a, reached a voltage- independent minimum at potentials positive to 0 mV. IA currents inactivated with two time constants that were voltage independent at potentials ranging from -30 to +45 mV. At +20 mV, tau i(fast) and tau i(slow) were 13.16 +/- 0.64 and 62.26 +/- 5.35 ms (n = 34), respectively. In some cells, IA inactivation kinetics slowed dramatically after many minutes of whole cell recording. Once activated by depolarization, IA channels returned to the closed state along pathways with two voltage-dependent time constants which were 0.208 s, tau rec-f and 10.02 s, tau rec-s at -80 mV. Approximately 90% of IA current recovered with slow kinetics at potentials between -60 and -100 mV. IA was blocked by 4-aminopyridine (IC50 = 629 microM) through a mechanism that was strongly promoted by channel activation. Divalent and trivalent cations including Ni2+ and La3+ also blocked IA with IC50's of 467 and 26.4 microM, respectively. With respect to biophysical properties and pharmacology, IA in AZF cells resembles to some extent transient K+ currents in neurons and muscle, where they function to regulate action potential frequency and duration. The function of this prominent current in steroid hormone secretion by endocrine cells that may not generate action potentials is not yet clear. 相似文献
13.
Pavlovian conditioning of the nudibranch mollusc Hermissenda crassicornis was previously shown to produce long-lasting reduction of two K+ currents measured across the Type B photoreceptor soma membrane (Alkon et al., 1982a; Alkon et al., 1985). Pavlovian conditioning of the rabbit was also shown to be followed by persistent K+ current reduction (Disterhoft et al., 1986). Here we report the first evidence that Ca2+ currents can also be modified by conditioning. The amplitude of the currents rather than their voltage-dependence remains reduced at least 1-2 d after conditioning (but not control procedures). Conditioning-induced changes of both K+ and Ca2+ currents increased as a function of training, the Ca2+ currents only changing substantially with greater than or equal to 250 trials. The later changes of the Ca2+ current may function to limit the magnitude of excitability increases due to associative learning. 相似文献
14.
Blackwell KT 《Journal of computational neuroscience》2004,17(1):81-99
The sea slug Hermissenda learns to associate light and hair cell stimulation, but not when the stimuli are temporally uncorrelated. Memory storage, which requires an elevation in calcium, occurs in the photoreceptors, which receive monosynaptic input from hair cells that sense acceleration stimuli such as turbulence. Both light and hair cell activity increase calcium concentration in the photoreceptor, but it is unknown whether paired calcium signals combine supralinearly to initiate memory storage. A correlate of memory storage is an enhancement of the long lasting depolarization (LLD) after light offset, which is attributed to a reduction in voltage dependent potassium currents; however, it is unclear what causes the LLD in the untrained animal.These issues were addressed using a multi-compartmental computer model of phototransduction, calcium dynamics, and ionic currents of the Hermissenda photoreceptor. Simulations of the interaction between light and hair cell activity show that paired stimuli do not produce a greater calcium increase than unpaired stimuli. This suggests that hair cell activity is acting via some other pathway to initiate memory storage. In addition, simulations show that a potassium leak channel, which closes with an increase in calcium, is required to produce both the untrained LLD and the enhanced LLD due to the decrease in voltage dependent potassium currents. Thus, the expression of this correlate of classical conditioning may depend on a leak potassium current. 相似文献
15.
Monoclonal antibodies to the p21 products of the transforming gene of Harvey murine sarcoma virus and of the cellular ras gene family. 总被引:47,自引:10,他引:37
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We have isolated eight rat lymphocyte-myeloma hybrid cell lines producing monoclonal antibodies that react with the 21,000-dalton transforming protein (p21) encoded by the v-ras gene of Harvey murine sarcoma virus (Ha-MuSV). These antibodies specifically immunoprecipitate both phosphorylated and non-phosphorylated forms of p21 from lysates of cells transformed by Ha-MuSV. All eight react with the products of closely related ras genes expressed in cells transformed by two additional sarcoma viruses (rat sarcoma virus and BALB sarcoma virus) or by a cellular Harvey-ras gene placed under the control of a viral promoter. Three of the antibodies also react strongly with the p21 encoded by the v-ras gene of Kirsten MuSV. These same three antibodies immunoprecipitate the predominant p21 species synthesized normally in a variety of rodent cell lines, including the p21 produced at high levels in 416B murine hemopoietic cells. This suggests that an endogenous gene closely related to Kirsten-ras is expressed in these cells. The monoclonal antibodies have been used to confirm two properties associated with p21; localization at the inner surface of the membrane of Ha-MuSV-transformed cells, assayed by immunofluorescence microscopy, and binding of guanine nucleotides. 相似文献
16.
Expression of voltage dependent potassium currents in freshly dissociated rat articular chondrocytes. 总被引:3,自引:0,他引:3
Arturo Ponce 《Cellular physiology and biochemistry》2006,18(1-3):35-46
The electrophysiological properties of voltage dependent potassium channels from freshly dissociated rat articular chondrocytes were studied. The resting membrane potential (-42.7+/-2.0 mV) was significantly depolarized by increasing concentrations of external potassium. No change was observed when external chloride concentration was varied. Addition of TEA, 4AP, alpha-Dendrotoxin and charybdotoxin depolarized resting membrane potential. Whole cell patch clamp studies revealed the presence of outwardly rectifying currents whose kinetic and pharmacological properties suggest the expression of voltage dependent potassium channels. Two kinds of currents were observed under the same experimental conditions. The first one, most frequently observed (80%), starts activating near -50 mV, with V(1/2)=-18 mV, G(max)=0.30 pS/pF. The second kind was observed in only 10% of cases; It activates near -40 mV, with(1/2)=+28.35 mV, G(max)=0.28 pS/pF pA/pF and does not inactivates. Inactivating currents were significantly inhibited by TEA (IC(50)=1.45 mM), 4AP (IC(50)=0.64 mM), CTX (IC(50) = 10 nM), alpha-Dendrotoxin (IC(50) < 100 nM) and Margatoxin (IC(50)=28.5 nM). These results show that rat chondrocytes express voltage dependent potassium currents and suggest a role of voltage-dependent potassium channels in regulating membrane potential of rat chondrocytes. 相似文献
17.
Serotonin decreases a background current and increases calcium and calcium-activated current in pedal neurons ofHermissenda 总被引:1,自引:0,他引:1
The effects of serotonin (5-HT) on membrane potential, membrane resistance, and select ionic currents were examined in large pedal neurons (LP1, LP3) of the mollusk Hermissenda. Calcium (Ca) action potentials were evoked in sodium-free artificial seawater containing tetramethylammonium, tetraethylammonium, and 4-aminopyridine (0-Na, 4-AP, TEA ASW). They failed at stimulation rates greater than 0.5/sec and were blocked by cadmium (Cd). Under voltage clamp the calcium current (ICa) responsible for them also failed with repeated stimulation. Thus, ICa inactivation accounts for refractoriness of the Ca action potential. The addition of 10 microM 5-HT to 0-Na, 4-AP, TEA ASW produced a slight depolarization and increased excitability and input resistance. Under voltage clamp the background current decreased. The voltage-dependent inward, late outward, and outward tail currents, sensitive to Cd, increased. ICa inactivation persisted. Under voltage clamp with Ca influx blocked by Cd, the addition of 10 microM 5-HT decreased the remaining current uniformly over membrane potentials of -10 to -100 mV. Thus, 5-HT reduces a background current that is active within the physiological range of the membrane potential, voltage insensitive, independent of Ca influx, noninactivating, and not blocked by 4-AP or TEA. 相似文献
18.
In mammalian systems, Ca2+/diacylglycerol-activated phospholipid-dependent protein kinase (C-kinase) appears to play an important role in regulating physiological responses that outlast the transient rise in cytosolic Ca2+. Electrophysiological experiments in neurons of the nudibranch mollusc, Hermissenda crassicornis, have suggested a role for C-kinase in the long-lasting reductions in early and late K+ currents that have been observed following associative learning. Accordingly, we have investigated the catalytic properties of C-kinase in Hermissenda CNS. Following homogenization in Ca2+-free buffer, C-kinase can be separated from Ca2+/calmodulin-dependent protein kinase by centrifugation; C-kinase activity is found in the supernatant whereas essentially all of the Ca2+/calmodulin-dependent protein kinase is found in the membrane fraction. Addition of Ca2+, phosphatidylserine, and diacylglycerol to the cytosol results in phosphorylation of at least eight endogenous proteins. The Hermissenda CNS C-kinase can also phosphorylate lysine-rich histone, a substrate for mammalian C-kinase. The molluscan enzyme exhibits phospholipid specificity in that phosphatidylserine is much more effective than phosphatidylethanolamine, phosphatidylcholine, phosphatidylinositol, and phosphatidic acid. Addition of diacylglycerol, in the presence of Ca2+ and phosphatidylserine, increases the activity of the C-kinase. The percentage of activation by diacylglycerol is larger at lower Ca2+ concentrations. Enzyme activity is inhibited by trifluoperazine and polymixin B sulfate. These studies indicate that the Hermissenda C-kinase is catalytically similar to mammalian C-kinase. 相似文献
19.
The effects of Pentylenetetrazol (PTZ) on the metacerebral giant cell (MCC) of the snail, Helix pomatia were studied. Actions on membrane resistance, time constant, resting and action potentials, outward and inward ionic currents were examined. Superfusion with PTZ in concentrations of 25 to 50 mmol/l, induced a gradually evolving convulsive state, which could be studied by intracellular recording from the MCCs. In the pre-convulsive state an acceleration of the spontaneous activity developed and was followed by paroxysmal depolarization shifts (PDSs), in the convulsive phase. PTZ prolonged the membrane time constant by about 10 percent, but this could not be traced back to alterations in membrane resistance or capacity. The resting membrane potential was not significantly altered; the action potentials were prolonged by slowing down of both the rising and decaying phases. The outward potassium currents were repressed by PTZ in a voltage dependent manner. The decrease of the IA current became more pronounced at increasingly positive command pulses, while IK was relieved from depression especially at longer pulse durations. Inward currents were isolated with the aid of suppression of outward currents by 50 mmol/l TEA. Under these conditions sodium currents, measured in calcium deficient Ringer solution were moderately depressed, while the calcium currents, examined during sodium-free superfusion, were mildly enhanced by PTZ. It is concluded that PTZ effects on ionic conductances, on membrane parameters, on the resting potential and ionic currents explain only modifications of spike potentials occurring in the convulsive state and do not account for the PDS, the central phenomenon of the convulsive electrographic activity, at least in this thoroughly examined type of neuron. 相似文献
20.
N Madi G Steiger J Estreicher J A Schifferli 《Journal of immunology (Baltimore, Md. : 1950)》1991,147(2):495-502
Mixed essential cryoglobulinemia type II (monoclonal Ig/polyclonal IgG) is characterized by systemic vasculitis caused by the deposition of circulating immune reactants that include the monoclonal component. Such reactants may include immune complexes (IC) formed from exogenous Ag. IC binding to E C receptor type 1 appears to play a role in transport and buffering of such IC (immune adherence: IA). To define the mechanisms responsible for immune deposition, 7 patients with cryoglobulinemia type II (IgM kappa/polyclonal IgG) and 14 normal volunteers were injected i.v. with hepatitis B surface Ag/antibody complexes. Two minutes after injection, only 19.4% (mean) of the circulating complexes were bound to E in patients as compared with 63.1% in normal subjects. This IA correlated directly with C4 and inversely with the IgM rheumatoid factor (RF) titer. Disappearance of IC was faster in patients (mean elimination rate: 15.7%/min) than in normal subjects (9.3%). In vitro experiments demonstrated that C depletion, interference with IC opsonization by monoclonal IgM RF, and decreased binding of opsonized IC in the presence of monoclonal RF are each associated with decreased IA. These observations suggest that, in patients with cryoglobulinemia type II, monoclonal IgM RF and low C contribute to reducing IA of circulating IC that might be rapidly trapped in tissues, resulting in injury. 相似文献