Biomedical subjects
Publications and source records attributed to M Fillenz.
Cyclic adenosine monophosphate (cAMP) concentration was measured in rat striatal and hippocampal synaptosomes. Incubation in 50 mM K+ medium caused a decrease in both striatal and hippocampal cAMP: this effect was dependent on extracellular Ca2+. Replacement of NaCl with sucrose in the incubation medium increased cAMP. Incubation with 0.5 mM ouabain was without effect.
The effects of alpha 2-adrenoceptor and gamma-aminobutyric acidB (GABAB) receptor activation on noradrenaline synthesis were measured in rat hippocampal synaptosomes. The results show that alpha 2-autoreceptors and GABAB heteroreceptors inhibit K+-accelerated synthesis; they have no effect on basal synthesis.
Rats were chronically implanted with recording electrodes in the dorsomedial subiculum and stimulating electrodes in the septal area. Low-frequency septal stimulation (regular inter-pulse interval = 130 msec) was used over a period of 10 days to drive the hippocampal theta rhythm at 7.7 Hz (a régime shown previously to proactively facilitate acquisition of barpressing and retard its extinction), and high frequency (77 Hz) stimulation was used to block theta rhythm (a régime shown previously to proactively retard acquisition of barpressing and facilitate its extinction). The activity of soluble tyrosine hydroxylase from hippocampus increased in animals killed 15-33 days after the end of the period of 7.7 Hz septal stimulation but not after high-frequency stimulation. Irregular low-frequency stimulation with a mean inter-pulse of 130 msec produced the same effect on tyrosine hydroxylase activity as regular 7.7 Hz theta-driving. The possible relations between the effects of low-frequency septal stimulation on hippocampal tyrosine hydroxylase and on behavior are discussed.
A procedure to measure noradrenaline synthesis in rat hippocampal synaptosomes using high-performance liquid chromatography with electrochemical detection is described. In order to characterize the technique, the linearity of synthesis as well as the effects of K+-depolarization and tyrosine omission were investigated. The technique was compared with the radiochemical tritium release method and the DOPA-accumulation method.
Changes in homovanillic acid concentration, recorded in vivo by voltammetry with carbon-paste electrodes, were used as an index of dopamine release. With electrodes implanted in frontal cortex, nucleus accumbens and striatum, the pattern of dopamine release was monitored simultaneously in the three brain regions together with the rats' motor activity. Changes in response to the systemic administration of dopamine-receptor agonists and antagonists were used as an index of feedback control of dopamine release. The relationships between dopamine release and motor activity, as well as that between dopamine release in the different brain regions, were investigated by calculating correlation coefficients for data collected over 24-h periods. The results show that dopamine release in frontal cortex is subject to little feedback regulation, that there is no nocturnal increase and no correlation with motor activity. Dopamine release in accumbens and striatum follows a very different pattern. There was a high correlation between dopamine release in these two regions on both sides of the brain; the correlation between the accumbens and the ipsilateral striatum was the highest. Dopamine release in both regions shows evidence of considerable feedback regulation, a nocturnal increase and a high correlation with motor activity. The importance of the accumbens in relation to the level of motor activity is supported by the finding that the correlation coefficient between motor activity and dopamine release in this structure was significantly higher than that between activity and release in the striatum.
Linear sweep voltammograms recorded with carbon paste electrodes in the striatum of the unanaesthetised, unrestrained rat show three separate peaks. The effect on peak 3 of either unilateral 6-hydroxydopamine lesion of the substantia nigra or intraperitoneal administration of alpha-methyl-paratyrosine, supports our earlier conclusion that peak 3 is due to the dopamine metabolite homovanillic acid. Administration of gamma-butyrolactone, which inhibits firing of dopaminergic nigrostriatal neurones, produces an immediate decrease in striatal homovanillic acid, followed by a prolonged increase. Dopamine-receptor agonists and antagonists produce changes in the extracellular concentration of homovanillic acid which are predicted by their effects on dopamine release. Simultaneous monitoring of total motor activity and homovanillic acid show significant correlation between these two parameters. The usefulness of this technique for monitoring dopamine release is critically evaluated in the light of these results.
The effects of guanyl nucleotides on the binding of [3H]flunitrazepam to rat hippocampal synaptic membranes were studied. In equilibrium binding studies, gamma-amino-n-butyric acid (GABA) increased and GTP decreased the binding affinity of [3H]flunitrazepam; GTP also caused a decrease in binding capacity. The effect, however, is variable. In studies of the dissociation kinetics of [3H]flunitrazepam using diazepam and the antagonist Ro 15-1788 as the displacers, there was evidence of two dissociation rate constants. GTP increased both the fast- and slow-dissociation rate constants and increased the ratio of the slow-dissociation binding state. The effect of GTP was mimicked by its nonhydrolyzable analogue 5'-guanylylimidodiphosphate but not by ATP and occurred when diazepam, but not when Ro 15-1788, was used as the displacer. GABA antagonized the effect of GTP on the dissociation of [3H]flunitrazepam. The nature of the benzodiazepine receptor, its actions, and the possible role of cyclic AMP as a second messenger are discussed.
To discover the significance of changes in the extracellular concentration of brain ascorbate, we used linear sweep voltammetry to monitor the ascorbate signal. Recordings were made with carbon paste electrodes implanted in the striatum and hippocampus of anaesthetised and unanaesthetised rats under a variety of conditions. Intraperitoneal administration of excitatory amino acid transmitters, but not tyrosine or glycine, increased extracellular striatal ascorbate; similarly, microinfusion of L-glutamate beside striatal electrodes enhanced the ascorbate signal. Electrical stimulation of the perforant path increased the extracellular concentration of dentate ascorbate in the unanaesthetised, but not in the anaesthetised, rat. These results support our hypothesis that changes in the extracellular concentration of brain ascorbate monitored by voltammetry reflect the release of excitatory amino acids.
The actions on [3H]noradrenaline release of chlordiazepoxide and FG 7142 were investigated in rat hippocampal synaptosomes. The release evoked by GABA, an action mediated by GABAA receptors, was enhanced by chlordiazepoxide and depressed by FG 7142. K+-evoked release, however, was depressed by both chlordiazepoxide and FG 7142 and occurred in the absence of GABA. The actions on both GABA-evoked and K+-evoked release were blocked by Ro 15-1788. The results suggest that the distinction between agonist and inverse agonist applies to the GABA-dependent but not to the GABA-independent action of benzodiazepines.
The effects of repeated mild stress on cortical alpha 2- and beta-adrenoceptor binding sites and on the rate of noradrenaline synthesis in cortical synaptosomes were measured. The stressful stimuli were either daily handling (1 min) or daily handling which included an i.p. saline injection. In both cases, rats killed 24 h after the last of 14 daily treatments showed an increase in noradrenaline synthesis rate and a decrease of alpha 2- and beta-adrenoceptor density. There were no changes in adrenoceptor density or the noradrenaline synthesis rate after either a single or 5 daily handlings plus saline injections. At no time was a change found in the Kd of either alpha 2- or beta-adrenoceptor binding sites. The results are discussed in relation to the effects on adrenoceptors of chronic antidepressant treatments.
Changes in the height of peak 2 obtained using linear sweep voltammetry and carbon paste electrodes chronically implanted in discrete brain regions of the unrestrained rat were measured under a variety of conditions; in the past this peak has been attributed to the oxidation of 5-hydroxyindoleacetic acid (5-HIAA). Unilateral 5,7-dihydroxytryptamine (5,7-DHT) lesions of the medial forebrain bundle reduced the 5-HIAA content of the striatum and hippocampus to 10% of the unlesioned side, but did not alter the height of peak 2 recorded in these regions. In contrast, microinfusion of uricase beside striatial electrodes reduced the height of peak 2 by 96%; systemic amphetamine-induced increases in the height of the peak were also prevented by this enzyme. These results indicate that uric acid, and not 5-HIAA, is mainly responsible for peak 2, and that changes in the height of this peak reflect changes in the extracellular concentration of uric acid.
Barbiturates are believed to work both by augmenting GABA action and independently perhaps by decreasing neurotransmitter release. By studying the effect of pentobarbitates on the release of 3H noradrenaline from rat hippocampal synaptosomes it was found that pentobarbitone (10(-6)M and 10(-5)M) augmented the GABA evoked release of 3H noradrenaline but 10(-4)M depressed this GABA effect. This latter concentration of pentobarbitone also depressed the K+-evoked release of 3H noradrenaline by a Ca++ dependent but picrotoxin insensitive mechanism.
Rats with neurotoxic lesions of the dorsal ascending noradrenergic bundle (DB) were compared with sham-operated (SH) controls on the acquisition, steady state and extinction of response suppression maintained by a classical (conditioned suppression) or an instrumental (discriminated punishment) contingency. DB lesions interfered neither with the acquisition of the reference response of sucrose-rewarded barpressing nor with unconditioned responding to the overhead flashing light subsequently used as a signal of shock. The acquisition of discriminated response suppression was also unaffected by the lesion under both types of contingency. However, once discriminated suppression had stabilized, both the conditioned and the discriminative stimulus used were significantly less effective in maintaining suppression in DB animals than in SH controls provided that low intensity footshock (0.2 mA) was used as the unconditioned stimulus (UCS). Upon increase of UCS intensity (to 0.5 mA) normal suppression was observed in the DB group under both contingencies. Extinction of the classical contingency reinstated the difference between DB and SH performance: DB lesion resulted in significantly faster extinction of fear. In contrast, extinction of the discriminated punishment contingency was unaffected by the lesion, although generalized response suppression dissipated faster in the DB than in the SH animals trained under this condition. Our results offer no support for the reinforcement hypothesis of DB function (normal acquisition of barpressing and of discriminated suppression of barpressing); mixed support (greater initial generalization of suppression in DB animals) and contradiction (more rapid extinction of conditioned suppression in DB animals) for the attentional hypothesis; and weak support (reduced suppression and more rapid extinction of suppression in DB animals, but only within limited experimental parameters) for the anxiety hypothesis of DB function. Hence none of the extant theories of DB function offer a ready explanation of the pattern of results presented here. A simple interpretation which conforms with the sparsity of positive behavioural findings in the literature on DB lesions is that forebrain noradrenaline contributes to the detection and utilization of conditioned stimuli; but that this contribution is critical only for the detection of stimuli with low associative strength.
Stimulation of the septal area at a frequency between 6 and 10 Hz is able to drive hippocampal theta. In freely moving male rats, the minimum threshold current for driving theta occurs at 7.7 Hz. Disruption of the pituitary-adrenal axis by injection of corticosterone to normal rats or by bilateral adrenalectomy (ADX) causes a shift of the minimum theta-driving threshold to 6.9 Hz. Corticosterone injection to ADX rats returns the minimum to 7.7 Hz. Specific and localized removal of hippocampal serotonergic fibers by intracerebral injections of 5,7-dihydroxytryptamine (5,7-DHT) produces the same shift to a 6.9-Hz minimum threshold as does corticosterone or ADX. We further report that these effects of manipulating the adrenocortical and serotonergic systems act through related (or common) mechanisms since: (1) 5,7-DHT shift to 6.9 Hz can be reversed to 7.7 Hz by injection of corticosterone; (2) 5,7-DHT lesions in an ADX rat produce a normal theta threshold minimum at 7.7 Hz, and (3) in a combined ADX- and 5,7-DHT-lesioned rat, corticosterone again produces a 6.9-Hz minimum as this hormone does in normal rats. These results suggest that the serotonergic inputs to the hippocampus interact with the same neurons which concentrate corticosterone.
Noradrenaline release from synaptosomes of rat hippocampus is modulated by both GABA and benzodiazepines. GABAA and GABAB receptors are present on these nerve terminals, and agonists at these receptors enhance spontaneous release and depress K+ -evoked release, respectively. Chlordiazepoxide has a dual effect: it enhances the action of GABA at GABAA receptors and depresses K+ -evoked release of [3H]noradrenaline in the absence of GABA. The mechanism of these actions and their in vivo role are discussed.
Linear sweep voltammograms were recorded in the striata of rats with a unilateral cortical lesion. The height of the ascorbate peak was 55% smaller on the lesioned side compared with the intact side, the homovanillic acid peak showed no significant change. The nocturnal increase in the release of striatal ascorbate was reduced by 80% on the lesioned side whilst the circadian variation in homovanillic acid was unaffected. These results support the hypothesis that there is a link between glutamate release and extracellular ascorbate concentrations in the brain.
Experimental properties of the peaks obtained with carbon paste electrodes and microprocessor-based linear sweep voltammetry in the rat brain have been compared with those predicted by a theoretical treatment of the problem which we have recently developed. This comparison indicates that the electrodes are situated in a restricted compartment and that the oxidation of ascorbate under these conditions is irreversible. Examples of how this situation has been exploited to improve the voltammograms obtained are given.
A number of advances have been made in the application of linear sweep voltammetry with carbon paste electrodes to the monitoring of ascorbate and monoamine neuroregulator metabolites in the brain of the freely-moving rat. These new measures are: the reduction of the sweep rate of 5 mV/s; the elimination of semidifferentiation in chronic recordings; the measurement in situ and subtraction of the background current for each electrode; the reduction, to practically zero levels, of the ascorbate signal in alternate scans; and corrections for circadian changes. These innovations enable one to monitor changes in the monoamine metabolites, even when there are simultaneous, large changes in ascorbate.