Peter Palij

Application of fast cyclic voltammetry to measurement of electrically evoked dopamine overflow from brain slices in vitro

Fast cyclic voltammetry at a carbon fibre microelectrode was used to monitor the time course of dopamine overflow in slices of rat corpus striatum incubated in a brain slice chamber. Dopamine release occurred in response to electrical stimulation. Electrochemical, physiological and pharmacological evidence indicates that release of endogenous dopamine can be measured reliably for up to 9 h and that fast cyclic voltammetry can be used in brain slices for quantitative studies of dopamine release in the CNS.

Presynaptic regulation of dopamine release in corpus striatum monitored in vitro in real time by fast cyclic voltammetry

Dopamine release was evoked by single electrical pulses in slices of rat corpus striatum, and measured by fast cyclic voltammetry in real time. The magnitude of the release varied in the expected way to agents which modify dopamine storage, release and re-uptake. The presence of functional dopamine D2 autoreceptors was demonstrated by showing that the release was potently and completely inhibited by the selective agonists quinpirole and N,N-dipropyl-5,6-ADTN. The selective D1 agonist SKF 38393 was ineffective. The inhibition by quinpirole was competitively antagonised by haloperidol and metoclopramide with potencies which correspond closely to published values at postsynaptic striatal D2 receptors. Thus, the D2 autoreceptors on striatal nerve terminals appear to be indistinguishable from those on the postsynaptic neurons.

Diffusion and uptake of dopamine in rat caudate and nucleus accumbens compared using fast cyclic voltammetry

Fast cyclic voltammetry was used in the caudate and nucleus accumbens of anaesthetised rats to study the release and reuptake of dopamine following stimulation of the median forebrain bundle. Dopamine uptake was significantly slower in accumbens than caudate, indicating a lower number of functional uptake sites. This implies that dopamine may be able to diffuse further from its sites of release in nucleus accumbens than in caudate and thus may have a neuromodulator role in this region.

Simultaneous “real-time” electrochemical and electrophysiological recording in brain slices with a single carbon-fibre microelectrode

Many previous studies have demonstrated the value of carbon-fibre microelectrodes (CFMs) for single-unit activity recording and for fast cyclic voltammetry. In this report we show that these two independent methodologies can be combined at a single CFM and used to study simultaneous electrochemical and electrophysiological events in brain slices. In superfused slices of rat locus coeruleus, dorsal raphe and substantia nigra, we were able to record stable electrophysiological signals and stimulated monoamine efflux for periods of at least 2 h, thereby allowing quantitative pharmacological interventions. The simultaneous recording of amine efflux and unit activity at the same locus facilitates comparison of drug effects at pre- and post-synaptic sites. Furthermore, the system described here uses commercially available instrumentation. The circuitry is described and examples of its application are shown.

Real-time monitoring of endogenous noradrenaline release in rat brain slices using fast cyclic voltammetry. 2. Operational characteristics of theα2 autoreceptor in the bed nucleus of stria terminalis, pars ventralis

Fast cyclic voltammetry (FCV) at carbon fibre microelectrodes was used to monitor stimulated noradrenaline (NA) efflux in slices of the ventral part of the rat bed nucleus of stria terminalis (BSTV) superfused with artificial cerebrospinal fluid at 32 degrees C. NA efflux was evoked by local electrical stimulation (trains of 10-50 pulses, 0.2 ms duration, 10 mA constant current at 10-500 Hz). The effects of four alpha 2 antagonists (yohimbine, rauwolscine, prazosin and WB 4101) and three alpha 2 agonists (clonidine, oxymetazoline and UK 14304) were examined. All drugs (1 microM) were added via the superfusate. Yohimbine and rauwolscine increased NA efflux on the lower but not the higher frequency trains: maximum increases (on 20 Hz, 50 pulse stimulation) were to 392 +/- 63% (yohimbine) and 243 +/- 7% (rauwolscine). There was a threshold train duration for demonstration of autoreceptor antagonism of 500-1000 ms. Prazosin and WB 4101 did not increase NA efflux but caused a modest decrease at the higher (100-500 Hz) frequencies. The effects of the alpha 2 agonists were also affected by stimulus train duration. Longer trains reduced agonist (clonidine) effects. When tested on pseudo-one pulse (POP) stimulations (less than 100 ms duration), the alpha 2 agonists decreased NA efflux. UK 14304 reduced NA efflux on 20 pulse/200 Hz stimulation to a greater degree (86 +/- 7%) than the partial agonists clonidine (39 +/- 3%) or oxymetazoline (40 +/- 8%). The present results demonstrate that alpha 2 autoreceptors are a major mechanism in the control of NA efflux in the BSTV.(ABSTRACT TRUNCATED AT 250 WORDS)

Biphasic inhibition of stimulated endogenous dopamine release by 7-OH-DPAT in slices of rat nucleus accumbens

1 Fast cyclic voltammetry was used to investigate the effect of 7‐OH‐DPAT (7‐hydroxy‐N,N‐di‐n‐propyl‐2‐aminotetralin), a putative D3 receptor agonist, on electrically stimulated endogenous dopamine release in slices of rat nucleus accumbens. 2 7‐OH‐DPAT inhibited single pulse stimulated dopamine release in a concentration‐dependent manner with a maximum inhibition of 95.5%. Analysis of concentration‐response curves to 7‐OH‐DPAT showed that they were biphasic, with the high affinity component contributing 18.0% to the total inhibition and the low affinity component 77.5%. 7‐OH‐DPAT exhibited a 560 fold selectivity between the high and low affinity components (0.015 nm compared to 8.4 nM). 3 Concentration‐response curves to the nonselective D2/D3 agonist, apomorphine, were monophasic. The maximum inhibition was 93.1% and the EC50 value 82 nM. 4 The selective D2 antagonist, haloperidol (30 nM), antagonized the low affinity component of the concentration‐response cuve to 7‐OH‐DPAT whilst the high affinity component was essentially unaffected. The pKB values calculated for the high and low affinity components were 7.89 and 9.45 respectively. 5 In conclusion, these results demonstrate that 7‐OH‐DPAT inhibits stimulated dopamine release by acting at two different sites. Furthermore, the results are consistent with the hypothesis that the high and low affinity components of the concentration‐response curve to 7‐OH‐DPAT may reflect activation of functional D3 and D2 release‐regulating autoreceptors respectively. However, the possibility that the biphasic nature of the curve may reflect different subtypes of the D2 receptor cannot be excluded.

Fast cyclic voltammetry: neurotransmitter measurement in “real time” and “real space”

Abstract In functional terms, the brain is highly spatially and temporally organised. Its function may be studied by many types of implantable probe such as dialysis fibres, cannulae or microelectrodes. In vivo voltammety, in particular, has been extensively used to study neurochemistry in situ. However, spatial and temporal sampling constraints can profoundly affect not only the reproducibility but also the accuracy of all neurochemical data obtained by voltammetry and related sampling techniques. For instance, it is possible to get inaccurate yet highly reproducible temporal information about neurotransmission simply by injudicious choice of sampling parameters. This review discusses the importance of voltammetric sampling parameters to the quality of neurochemical data that we, as neuroscientists, obtain. Although these considerations apply equally to all sampling based methodologies we have chosen to focus on the case of fast cyclic voltammetry. The review is illustrated by data using fast cyclic voltammetry at carbon fibre microelectrodes in brain slices.

Rauwolscine potentiates the effect of desipramine on limbic noradrenaline efflux.

DESIPRAMINE, like most antidepressants, does not exert clinical benefit until 2–3 weeks after the onset of treatment. It has been suggested that this delay might be due to enhanced autoreceptor activation, counteracting the acute uptake blockade. We therefore tested whether autoreceptor blockade might enhance the response to acute uptake blockade, using voltammetry at carbon fibre microelectrodes to monitor stimulated noradrenaline (NA) efflux in rat bed nucleus of stria terminalis brain slices. Desipramine significantly increased NA efflux and slowed NA uptake. The combination of rauwolscine and desipramine increased NA efflux significantly more than desipramine alone. We suggest that α2 autoreceptor blockade functionally mimics α2 autoreceptor down-regulation and thus may allow the full therapeutic effect of desipramine to be manifested more rapidly.