Long-term effects of N-2-chlorethyl-N-ethyl-2-bromobenzylamine hydrochloride on noradrenergic neurones in the rat brain and heart.
1 N-2-Chlorethyl-N-ethyl-2-bromobenzylamine hydrochloride (DSP 4) 50 mg/kg intraperitoneally, produced a long-term decrease in the capacity of brain homogenates to accumulate noradrenaline with significant effect 8 months after the injection. It had no effect on the noradrenaline uptake in homogenates from the striatum (dopamine neurones) and on the uptake of 5-hydroxytryptamine (5-HT) in various brain regions. 2 In vitro DSP 4 inhibited the noradrenaline uptake in a cortical homogenate with an IC50 value of 2 muM but was more than ten times less active on the dopamine uptake in a striatal homogenate and the 5-HT uptake in a cortical homogenate. 3 DSP 4 (50 mg/kg i.p.) inhibited the uptake of noradrenaline in the rat heart atrium in vitro but this action was terminated within 2 weeks. 4 DSP 4 (50 mg/kg i.p.) cuased a decrease in the dopamine-beta-hydroxylase (DBH) activity in the rat brain and heart. The onset of this effect was slow; in heart a lag period of 2-4 days was noted. In brain the DBH-activity in cerebral cortex was much more decreased than that in hypothalamus which was only slightly affected. A significant effect was still found 8 months after the injection. The noradrenaline concentration in the brain was greatly decreased for at least two weeks, whereas noradrenaline in heart was only temporarily reduced. 5 The long-term effects of DSP 4 on the noradrenaline accumulation, the DBH activity and noradrenaline concentration in the rat brain were antagonized by desipramine (10 mg/kg i.p.). 6 It is suggested that DSP 4 primarily attacks the membranal noradrenaline uptake sites forming a covalent bond and that the nerve terminals, as a result of this binding, degenerate. (+info
Dopamine stimulates salivary duct cells in the cockroach Periplaneta americana.
This study examines whether the salivary duct cells of the cockroach Periplaneta americana can be stimulated by the neurotransmitters dopamine and serotonin. We have carried out digital Ca2+-imaging experiments using the Ca2+-sensitive dye fura-2 and conventional intracellular recordings from isolated salivary glands. Dopamine evokes a slow, almost tonic, and reversible dose-dependent elevation in [Ca2+]i in the duct cells. Upon stimulation with 10(-)6 mol l-1 dopamine, [Ca2+]i rises from 48+/-4 nmol l-1 to 311+/-43 nmol l-1 (mean +/- s.e.m., N=18) within 200-300 s. The dopamine-induced elevation in [Ca2+]i is absent in Ca2+-free saline and is blocked by 10(-)4 mol l-1 La3+, indicating that dopamine induces an influx of Ca2+ across the basolateral membrane of the duct cells. Stimulation with 10(-)6 mol l-1 dopamine causes the basolateral membrane to depolarize from -67+/-1 to -41+/-2 mV (N=10). This depolarization is also blocked by La3+ and is abolished when Na+ in the bath solution is reduced to 10 mmol l-1. Serotonin affects neither [Ca2+]i nor the basolateral membrane potential of the duct cells. These data indicate that the neurotransmitter dopamine, which has previously been shown to stimulate fluid secretion from the glands, also stimulates the salivary duct cells, suggesting that dopamine controls their most probable function, the modification of primary saliva. (+info
Induction of serotonin transporter by hypoxia in pulmonary vascular smooth muscle cells. Relationship with the mitogenic action of serotonin.
-The increased delivery of serotonin (5-hydroxytryptamine, 5-HT) to the lung aggravates the development of hypoxia-induced pulmonary hypertension in rats, possibly through stimulation of the proliferation of pulmonary artery smooth muscle cells (PA-SMCs). In cultured rat PA-SMCs, 5-HT (10(-8) to 10(-6) mol/L) induced DNA synthesis and potentiated the mitogenic effect of platelet-derived growth factor-BB (10 ng/mL). This effect was dependent on the 5-HT transporter (5-HTT), since it was prevented by the 5-HTT inhibitors fluoxetine (10(-6) mol/L) and paroxetine (10(-7) mol/L), but it was unaltered by ketanserin (10(-6) mol/L), a 5-HT2A receptor antagonist. In PA-SMCs exposed to hypoxia, the levels of 5-HTT mRNA (measured by competitive reverse transcriptase-polymerase chain reaction) increased by 240% within 2 hours, followed by a 3-fold increase in the uptake of [3H]5-HT at 24 hours. Cotransfection of the cells with a construct of human 5-HTT promoter-luciferase gene reporter and of pCMV-beta-galactosidase gene allowed the demonstration that exposure of cells to hypoxia produced a 5.5-fold increase in luciferase activity, with no change in beta-galactosidase activity. The increased expression of 5-HTT in hypoxic cells was associated with a greater mitogenic response to 5-HT (10(-8) to 10(-6) mol/L) in the absence as well as in the presence of platelet-derived growth factor-BB. 5-HTT expression assessed by quantitative reverse transcriptase-polymerase chain reaction and in situ hybridization in the lungs was found to predominate in the media of pulmonary artery, in which a marked increase was noted in rats that had been exposed to hypoxia for 15 days. These data show that in vitro and in vivo exposure to hypoxia induces, via a transcriptional mechanism, 5-HTT expression in PA-SMCs, and that this effect contributes to the stimulatory action of 5-HT on PA-SMC proliferation. In vivo expression of 5-HTT by PA-SMC may play a key role in serotonin-mediated pulmonary vascular remodeling. (+info
Activity-dependent metaplasticity of inhibitory and excitatory synaptic transmission in the lamprey spinal cord locomotor network.
Paired intracellular recordings have been used to examine the activity-dependent plasticity and neuromodulator-induced metaplasticity of synaptic inputs from identified inhibitory and excitatory interneurons in the lamprey spinal cord. Trains of spikes at 5-20 Hz were used to mimic the frequency of spiking that occurs in network interneurons during NMDA or brainstem-evoked locomotor activity. Inputs from inhibitory and excitatory interneurons exhibited similar activity-dependent changes, with synaptic depression developing during the spike train. The level of depression reached was greater with lower stimulation frequencies. Significant activity-dependent depression of inputs from excitatory interneurons and inhibitory crossed caudal interneurons, which are central elements in the patterning of network activity, usually developed between the fifth and tenth spikes in the train. Because these interneurons typically fire bursts of up to five spikes during locomotor activity, this activity-dependent plasticity will presumably not contribute to the patterning of network activity. However, in the presence of the neuromodulators substance P and 5-HT, significant activity-dependent metaplasticity of these inputs developed over the first five spikes in the train. Substance P induced significant activity-dependent depression of inhibitory but potentiation of excitatory interneuron inputs, whereas 5-HT induced significant activity-dependent potentiation of both inhibitory and excitatory interneuron inputs. Because these metaplastic effects are consistent with the substance P and 5-HT-induced modulation of the network output, activity-dependent metaplasticity could be a potential mechanism underlying the coordination and modulation of rhythmic network activity. (+info
Mechanisms for generating the autonomous cAMP-dependent protein kinase required for long-term facilitation in Aplysia.
The formation of a persistently active cAMP-dependent protein kinase (PKA) is critical for establishing long-term synaptic facilitation (LTF) in Aplysia. The injection of bovine catalytic (C) subunits into sensory neurons is sufficient to produce protein synthesis-dependent LTF. Early in the LTF induced by serotonin (5-HT), an autonomous PKA is generated through the ubiquitin-proteasome-mediated proteolysis of regulatory (R) subunits. The degradation of R occurs during an early time window and appears to be a key function of proteasomes in LTF. Lactacystin, a specific proteasome inhibitor, blocks the facilitation induced by 5-HT, and this block is rescued by injecting C subunits. R is degraded through an allosteric mechanism requiring an elevation of cAMP coincident with the induction of a ubiquitin carboxy-terminal hydrolase. (+info
Ethanol exposure differentially alters central monoamine neurotransmission in alcohol-preferring versus -nonpreferring rats.
Individual differences in ethanol preference may be linked to differences in the functional activity of forebrain monoamine systems or their sensitivity to modification by ethanol. To test this hypothesis, basal extracellular concentrations of dopamine (DA) and serotonin (5-HT) in the nucleus accumbens as well as the effects of repeated ethanol pretreatment on the basal release of these transmitters were examined in alcohol-preferring (P), alcohol-nonpreferring (NP), and genetically heterogeneous Wistar rats. All animals received i.p. injections of ethanol (1.0 g/kg) or saline for 5 consecutive days. Fifteen hours after the final pretreatment, basal extracellular concentrations and "in vivo extraction fraction" values for DA and 5-HT were determined by no-net-flux in vivo microdialysis. In ethanol-naive rats, significant line differences were observed with high basal 5-HT release in P rats, low 5-HT release in NP rats, and intermediate 5-HT levels in Wistar rats. No differences among groups were noted in basal DA release. Ethanol pretreatment decreased basal extracellular 5-HT levels in P rats whereas increasing 5-HT efflux was seen in the Wistar and NP lines. In addition, ethanol pretreatment increased extracellular DA concentrations in Wistar and P rats, but not in NP rats. The results confirm a relationship between the functional status of forebrain DA and 5-HT systems and ethanol preference or aversion. Moreover, the data suggest that ethanol exposure can alter basal DA and 5-HT in the nucleus accumbens and that vulnerability to ethanol-induced changes in monoamine neurotransmission may be a factor in genetically determined ethanol preference. (+info
Possible role of serotonin in Merkel-like basal cells of the taste buds of the frog, Rana nigromaculata.
Merkel-like basal cells in the taste buds of the frog were examined by fluorescence histochemistry, immunohistochemistry and electron microscopy. There were about 16-20 basal cells arranged in a radial fashion at the base of each taste bud. These cells were strongly immunopositive for serotonin antiserum. They were characterised by the presence of numerous dense-cored granules in the cytoplasm ranging from 80 to 120 nm in diameter, and of microvilli protruding from the cell surface. For 4 mo after sensory denervation by cutting the gustatory nerves, all cell types of the taste bud were well preserved and maintained their fine structure. Even at 4 mo after denervation, the basal cells exhibited a strong immunoreaction with serotonin antiserum. To investigate the function of serotonin in the basal cells in taste bud function, serotonin deficiency was induced by administration of p-chlorophenylalanine (PCPA), an inhibitor of tryptophan hydroxylase, and of p-chloroamphetamine (PCA), a depletor of serotonin. After administration of these agents to normal and denervated frogs for 2 wk, a marked decrease, or complete absence, of immunoreactivity for serotonin was observed in the basal cells. Ultrastructurally, degenerative changes were observed in both types of frog; numerous lysosome-like myelin bodies were found in all cell types of the taste buds. The number of dense-cored granules in the basal cells also was greatly decreased by treatment with these drugs. Serotonin in Merkel-like basal cells appears to have a trophic role in maintenance of the morphological integrity of frog taste bud cells. (+info
Activation of stimulus-specific serine esterases (proteases) in the initiation of platelet secretion. I. Demonstration with organophosphorus inhibitors.
The effect of organophosphorus inhibitors of serine esterases (proteases) on secretion from washed rabbit platelets was examined. Five noncytotoxic stimuli were employed: collagen, thrombin, heterologous anti-platelet antibody (in the absence of complement), rabbit C3 bound to zymosan, and platelet activating factor derived from antigen-stimulated, IgE-sensitized rabbit basophils. Diisoprophyl phosphofluoridate, three series of p-nitrophenyl ethyl phosphonates, and a series of cyclohexyl phenylalkylphosphonofluridates were all found to be inhibitory to the platelet secretion. These are irreversible inhibitors of serine proteases but in this system were only inhibitory if added to the platelets concurrently with the stimuli. Pretreatment of either the platelets or the stimuli with the inhibitors followed by washing, was without effect on the subsequent reaction. This suggested the involvement of stimulus-activatable serine proteases in the secretory process. The concept was supported by finding that nonphosphorylating phosphonates or hydrolyzed phosphonates or phosphonofluoridates were without inhibitory action. The effect of a series of phosphonates or phosphonoflouridates in inhibiting each stimulus exhibited a unique activity-structure profile. The demonstration of such unique profiles with four series of inhibitors for each of the five stimuli was interpreted as demonstrating that a specific activatable serine protease was involved in the platelet secretory response to each stimulus. (+info