Electric Field Stimulation for the Functional Assessment of Isolated Dorsal Root Ganglion Neuron Excitability.

Genetically encoded calcium indicators have proven useful for characterizing dorsal root ganglion neuron excitability in vivo. Challenges persist in achieving high spatial-temporal resolutions in vivo, however, due to deep tissue imaging and motion artifacts that may be limiting technical factors in obtaining measurements. Here we report an ex vivo imaging method, using a peripheral neuron-specific Advillin-GCaMP mouse line and electric field stimulation of dorsal root ganglion tissues, to assess the sensitivity of neurons en bloc. The described method rapidly characterizes Ca2+ activity in hundreds of dorsal root ganglion neurons (221 ± 64 per dorsal root ganglion) with minimal perturbation to the in situ soma environment. We further validate the method for use as a drug screening platform with the voltage-gated sodium channel inhibitor, tetrodotoxin. Drug treatment led to decreased evoked Ca2+ activity; half-maximal response voltage (EV50) increased from 13.4 V in untreated tissues to 21.2, 23.3, 51.5 (p < 0.05), and 60.6 V (p < 0.05) at 0.01, 0.1, 1, and 10 µM doses, respectively. This technique may help improve an understanding of neural signaling while retaining tissue structural organization and serves as a tool for the rapid ex vivo recording and assessment of neural activity.

Assessment of TTX-s and TTX-r Action Potential Conduction along Neurites of NGF and GDNF Cultured Porcine DRG Somata.

Nine isoforms of voltage-gated sodium channels (NaV) have been characterized and in excitable tissues they are responsible for the initiation and conduction of action potentials. For primary afferent neurons residing in dorsal root ganglia (DRG), individual neurons may express multiple NaV isoforms extending the neuron's functional capabilities. Since expression of NaV isoforms can be differentially regulated by neurotrophic factors we have examined the functional consequences of exposure to either nerve growth factor (NGF) or glial cell line-derived neurotrophic factor (GDNF) on action potential conduction in outgrowing cultured porcine neurites of DRG neurons. Calcium signals were recorded using the exogenous intensity based calcium indicator Fluo-8®, AM. In 94 neurons, calcium signals were conducted along neurites in response to electrical stimulation of the soma. At an image acquisition rate of 25 Hz it was possible to discern calcium transients in response to individual electrical stimuli. The peak amplitude of electrically-evoked calcium signals was limited by the ability of the neuron to follow the stimulus frequency. The stimulus frequency required to evoke a half-maximal calcium response was approximately 3 Hz at room temperature. In 13 of 14 (93%) NGF-responsive neurites, TTX-r NaV isoforms alone were sufficient to support propagated signals. In contrast, calcium signals mediated by TTX-r NaVs were evident in only 4 of 11 (36%) neurites from somata cultured in GDNF. This establishes a basis for assessing action potential signaling using calcium imaging techniques in individual cultured neurites and suggests that, in the pig, afferent nociceptor classes relying on the functional properties of TTX-r NaV isoforms, such as cold-nociceptors, most probably derive from NGF-responsive DRG neurons.

Transient sodium current at subthreshold voltages: activation by EPSP waveforms.

Tetrodotoxin (TTX)-sensitive sodium channels carry large transient currents during action potentials and also "persistent" sodium current, a noninactivating TTX-sensitive current present at subthreshold voltages. We examined gating of subthreshold sodium current in dissociated cerebellar Purkinje neurons and hippocampal CA1 neurons, studied at 37°C with near-physiological ionic conditions. Unexpectedly, in both cell types small voltage steps at subthreshold voltages activated a substantial component of transient sodium current as well as persistent current. Subthreshold EPSP-like waveforms also activated a large component of transient sodium current, but IPSP-like waveforms engaged primarily persistent sodium current with only a small additional transient component. Activation of transient as well as persistent sodium current at subthreshold voltages produces amplification of EPSPs that is sensitive to the rate of depolarization and can help account for the dependence of spike threshold on depolarization rate, as previously observed in vivo.

Synaptic vesicle retrieval time is a cell-wide rather than individual-synapse property.

Although individual nerve terminals from the same neuron often differ in neurotransmitter release characteristics, the extent to which endocytic retrieval of synaptic vesicle components differs is unknown. We used high-fidelity optical recordings to undertake a large-scale analysis of endocytosis kinetics of individual boutons in hippocampal rat neurons. Our data indicate that endocytosis kinetics do not differ substantially across boutons from the same cell but instead appear to be controlled at a cell-wide level.

Up-regulation of tetrodotoxin-sensitive sodium currents by prostaglandin E2 in type-4 rat dorsal root ganglion cells.

Mechanisms were studied by which prostaglandin E(2) (PGE(2)) up-regulates Na(+) currents (INa) in medium diameter dorsal root ganglion (DRG) cells that express large T-type Ca(2+) currents (type-4 DRG cells). PGE(2) or the adenylyl cyclase (AC) activator forskolin (10 µM) up-regulated peak INa evoked by test potentials (TP) to -10 mV by an average of 13.5% and 21.8%, respectively. The PGE(2) and forskolin induced up-regulation of INa, evoked with TPs to -10 mV, began approximately 15-20 s after initiation of drug exposure and continued gradually over the course of 2-3 min. Both PGE(2) and forskolin significantly increased peak conductance without significantly shifting the voltage at which INa was ½ activated (V(a)) or ½ steady state inactivated. However, although V(a) was not significantly shifted, both PGE(2) and forskolin induced a proportionally greater percent increase in conductance at weak TPs to around -30 mV compared to stronger TPs to around 10 mV. The PGE(2)-induced up-regulation of INa was occluded by prior up-regulation with forskolin, and the up-regulation of INa by both PGE(2) and forskolin was blocked by Rp-cAMPs and 50 nM tetrodotoxin (TTX). In the presence of Rp-cAMPs, both PGE(2) and forskolin induced decreases in INa that peaked around 25 s following initiation of PGE(2)/forskolin application. The decrease induced by PGE(2) averaged 8.5%, which was significantly greater than the average 3.5% decrease induced by forskolin. Estimation of kinetic rate constants by fitting INa with a Markov channel state model, suggested that both PGE(2) and forskolin up-regulated INa by changing channel gating rather than by increasing channel number or unitary conductance. The data suggest that application of PGE(2) may initially induce a relatively rapid down-regulation of TTX-sensitive INa (signaling pathway uncharacterized), followed by a gradual up-regulation of INa via activation of an AC/PKA-dependent signaling pathway. The up-regulation of INa in sensory neurons with type-4 cell bodies may increase excitability and strengthen signaling, and may play some role in the allodynia and hyperalgesia associated with injury to nerves and peripheral tissues.

Biophysical costs associated with tetrodotoxin resistance in the sodium channel pore of the garter snake, Thamnophis sirtalis.

Tetrodotoxin (TTX) is a potent toxin that specifically binds to voltage-gated sodium channels (NaV). TTX binding physically blocks the flow of sodium ions through NaV, thereby preventing action potential generation and propagation. TTX has different binding affinities for different NaV isoforms. These differences are imparted by amino acid substitutions in positions within, or proximal to, the TTX-binding site in the channel pore. These substitutions confer TTX-resistance to a variety of species. The garter snake Thamnophis sirtalis has evolved TTX-resistance over the course of an arms race, allowing some populations of snakes to feed on tetrodotoxic newts, including Taricha granulosa. Different populations of the garter snake have different degrees of TTX-resistance, which is closely related to the number of amino acid substitutions. We tested the biophysical properties and ion selectivity of NaV of three garter snake populations from Bear Lake, Idaho; Warrenton, Oregon; and Willow Creek, California. We observed changes in gating properties of TTX-resistant (TTXr) NaV. In addition, ion selectivity of TTXr NaV was significantly different from that of TTX-sensitive NaV. These results suggest TTX-resistance comes at a cost to performance caused by changes in the biophysical properties and ion selectivity of TTXr NaV.

Cardiac basal metabolism: energetic cost of calcium withdrawal in the adult rat heart.

AIM: Cardiac basal metabolism upon extracellular calcium removal and its relationship with intracellular sodium and calcium homeostasis was evaluated. METHODS: A mechano-calorimetric technique was used that allowed the simultaneous and continuous measurement of both heat rate and resting pressure in arterially perfused quiescent adult rat hearts. Using pharmacological tools, the possible underlying mechanisms related to sodium and calcium movements were investigated. RESULTS: Resting heat rate (expressed in mW g(-1)(dry wt)) increased upon calcium withdrawal (+4.4 +/- 0.2). This response was: (1) unaffected by the presence of tetrodotoxin (+4.3 +/- 0.6), (2) fully blocked by both, the decrease in extracellular sodium concentration and the increase in extracellular magnesium concentration, (3) partially blocked by the presence of either nifedipine (+2.8 +/- 0.4), KB-R7943 (KBR; +2.5 +/- 0.2), clonazepam (CLO; +3.1 +/- 0.3) or EGTA (+1.9 +/- 0.3). The steady heat rate under Ca(2+)-free conditions was partially reduced by the addition of Ru360 (-1.1 +/- 0.2) but not CLO in the presence of EGTA, KBR or Ru360. CONCLUSION: Energy expenditure for resting state maintenance upon calcium withdrawal depends on the intracellular rise in both sodium and calcium. Our data are consistent with a mitochondrial Ca(2+) cycling, not detectable under normal calcium diastolic levels. The experimental condition here analysed, partially simulates findings reported under certain pathological situations including heart failure in which mildly increased levels of both diastolic sodium and calcium have also been found. Therefore, under such pathological conditions, hearts should distract chemical energy to fuel processes associated with sodium and calcium handling, making more expensive the maintenance of their functions.

Quantitative assessment of the distributions of membrane conductances involved in action potential backpropagation along basal dendrites.

Basal dendrites of prefrontal cortical neurons receive strong synaptic drive from recurrent excitatory synaptic inputs. Synaptic integration within basal dendrites is therefore likely to play an important role in cortical information processing. Both synaptic integration and synaptic plasticity depend crucially on dendritic membrane excitability and the backpropagation of action potentials. We carried out multisite voltage-sensitive dye imaging of membrane potential transients from thin basal branches of prefrontal cortical pyramidal neurons before and after application of channel blockers. We found that backpropagating action potentials (bAPs) are predominantly controlled by voltage-gated sodium and A-type potassium channels. In contrast, pharmacologically blocking the delayed rectifier potassium, voltage-gated calcium, or I(h) conductance had little effect on dendritic AP propagation. Optically recorded bAP waveforms were quantified and multicompartmental modeling was used to link the observed behavior with the underlying biophysical properties. The best-fit model included a nonuniform sodium channel distribution with decreasing conductance with distance from the soma, together with a nonuniform (increasing) A-type potassium conductance. AP amplitudes decline with distance in this model, but to a lesser extent than previously thought. We used this model to explore the mechanisms underlying two sets of published data involving high-frequency trains of APs and the local generation of sodium spikelets. We also explored the conditions under which I(A) down-regulation would produce branch strength potentiation in the proposed model. Finally, we discuss the hypothesis that a fraction of basal branches may have different membrane properties compared with sister branches in the same dendritic tree.

A novel toxicity-based assay for the identification of modulators of voltage-gated Na+ channels.

Voltage-gated Na+ channels are promising drug targets. Screening of large numbers of putative modulators, however, can be demanding and expensive. In this study, a simple, cheap, and robust assay to test the pharmacological modulation of Na+ channel function is presented. The assay makes use of the fact that the intracellular accumulation of Na+ ions can be cytotoxic. The toxicity of the Na+ channel activator veratridine in the presence of an inhibitor of the Na+/K+ ATPase (ouabain) in a Nav1.2a (rat brain IIA alpha) expressing cell line is assessed. Na+ channel blockers should reduce toxicity in this model. CHO cells which recombinantly expressed rat Nav1.2a subunits were seeded in 96-well plates, and cell survival was tested after 24 h incubation in medium containing veratridine and ouabain in the presence or absence of Na+ channel blockers. Propidium iodide fluorescence was used as toxicity readout. Veratridine (100 microM) or ouabain alone (500 microM) were not toxic to the cells. In the presence of 500 microM ouabain, however, veratridine induced halfmaximal cell death with an EC50 value of 15.1 +/- 2.3 microM. Ouabain's EC50 was 215.3 +/- 16.7 microM (with 30 microM veratridine). The effects of a number of Na+ channel blockers were tested and compared with their Na+ channel blocking activity measured in voltage-clamp experiments. Blockers from various chemical classes reduced toxicity half maximally with IC50 values ranging from 11.7 +/- 1.4 nM (tetrodotoxin) to 280.5 +/- 48.0 microM (lamotrigine). There was a linear relationship between the log IC50 values obtained by the two methods (slope: 1.1 +/- 0.08; correlation coefficient: 0.93). In summary, these data show that this novel toxicity assay is well suited to test Na+ channel blockers.

Resistance of neonates and field-collected garter snakes (Thamnophis spp.) to tetrodotoxin.

Prior studies of tetrodotoxin (TTX) resistance in garter snakes (Thamnophis spp.) have used laboratory-reared neonates as subjects, but the use of field-caught individuals would reduce cost and effort. We compared estimates of TTX resistance in field-caught and laboratory-born garter snakes. We found that a mass-adjusted dose of TTX administered to field-caught garter snakes produces an estimate of a population 50% dose that is comparable and unbiased with respect to those previously reported using laboratory-born neonates. Dose-response curves estimated for three field-caught populations closely matched the curves estimated from neonate data. The method was tested using populations with levels of TTX resistance ranging between approximately 5-90 mass-adjusted mouse units for their respective 50% doses. The technique of using field-caught snakes as test subjects provides larger genetically independent data sets that are more easily obtained. Our results indicate that changes in mass during development parallel ontogenetic shifts in TTX resistance.

In vivo electrical stimulation of rabbit retina with a microfabricated array: strategies to maximize responses for prospective assessment of stimulus efficacy and biocompatibility.

PURPOSE: Our primary goal was to assess the effects of varying stimulus parameters on the electrically evoked cortical potentials (EECPs) in rabbits, which we intend to use as one measure of biocompatibility of implanted retinal prosthetic devices. We also sought to exclude contamination of waveforms recorded over the occipital cortex from electrical activity from the retina and the degree of reproducibility of EECP recordings. METHODS: A concentric bipolar platinum electrode or microfabricated 5x5 electrode array delivered current to the retina of 43 Dutch-belted rabbits while the EECP was recorded from extradural electrodes over the occipital cortex. Electroretinogram (ERG) and visual evoked cortical potential (VECP) recordings were routinely obtained. Verification that occipital cortical recordings were not heavily contaminated by electrical potentials from the retina (i.e. the "validity" of the cortical recordings) was made by recording retinal and brain responses before and after intravitreal injection of tetrodotoxin. Electrical stimulation of the retina was performed with monopolar (with distant return) or bipolar electrode configurations. Cortical responses were computer-averaged over 100-500 stimulations. The effect of variation in stimulus current, charge, duration, frequency, polarity and spatial orientation of stimulating electrodes on cortical responses was studied. RESULTS: Progressive reduction of responses toward the anterior skull and abolition of posterior recordings by tetrodotoxin indicated that retinal activity did not significantly contaminate EECP recordings. Reproducibility testing revealed that inter-animal variability within the first hour of testing across all animals was not significantly greater than that found during prolonged testing of a single animal. The lowest current that yielded a reproducible EECP with monopolar stimulation was 75 microA (total current through 21 electrodes) using 200 microsec pulses, which yielded a 45 microV cortical response. Strength-duration curves were generally flat for fixed charge stimulation and linear for fixed current stimulation, at least up to a saturation point, which occurred at very high charge. Over 0.5-16 Hz stimulus frequencies, ERGs varied little but evoked potential responses showed a monotonic decline in amplitude at higher frequencies. Large negative-going initial pulses of a biphasic pair yielded the largest cortical amplitudes. EECP amplitudes varied significantly with the orientation of stimulating electrodes on the retina. CONCLUSIONS: This study provides novel data on the reproducibility of EECP recordings, and insight into stimulation parameters that affect retinal and cortical responses. This information can be used to improve the yield of retinal and evoked potential recordings, which will enhance the prospective assessment of the efficacy of stimulation and health of the stimulated tissues following.

Neuronal activity regulates correlated network properties of spontaneous calcium transients in astrocytes in situ.

Spontaneous neuronal activity is essential to neural development. Until recently, neurons were believed to be the only excitable cells to display spontaneous activity. However, cultured astrocytes and, more recently, astrocytes in situ are now known to exhibit spontaneous Ca2+ transients. Here we used Ca2+ imaging of astrocytes from transgenic mice for the simultaneous monitoring of [Ca2+]i changes in large numbers of astrocytes. We found that spontaneous activity is a common property of most brain astrocytes that is lost in response to a lesion. These spontaneous [Ca2+]i oscillations require extracellular and intracellular Ca2+. Moreover, network analysis revealed that most astrocytes formed correlated networks of dozens of these cells, which were synchronous with both astrocytes and neurons. We found that decreasing spontaneous [Ca2+]i transients in neurons by TTX does not alter the number of active astrocytes, although it impairs their synchronous network activity. Conversely, bicuculline-induced epileptic patterns of [Ca2+]i transients in neurons cause an increase in the number of active astrocytes and in their network synchrony. Furthermore, activation of non-NMDA and NMDA ionotropic glutamate receptors is required to correlate astrocytic networks. These results show that spontaneous activity in astrocytes and neurons is patterned into correlated neuronal/astrocytic networks in which neuronal activity regulates the network properties of astrocytes. This network activity may be essential for neural development and synaptic plasticity.

Feeding increases 5-hydroxytryptamine and norepinephrine within the hypothalamus of chicks.

It is thought that hypothalamic 5-hydroxytryptamine (5HT) and norepinephrine (NE) are involved in the regulation of feeding in chicks. The present study was conducted to elucidate changes in the levels of extracellular 5HT and NE in the hypothalamus during feeding of chicks. In order to measure 5HT, NE and 4-hydroxy-3-methoxyphenylglycol (MHPG), which is a major metabolite of NE, we used brain microdialysis and high-pressure liquid chromatography with an electrochemical detector. After collecting samples to determine the basal levels of 5HT, NE and MHPG, food-deprived birds were given access to food. 5HT levels in the medial hypothalamus (MH) and lateral hypothalamus (LH) increased during the first 30 min of feeding, and then returned to basal levels. NE and MHPG in the LH increased during feeding, and remained elevated throughout the experiment. This study supports an idea that hypothalamic monoamines in the chick brain are involved in the regulation of feeding.

In vivo assessment of the regulatory mechanism of cholinergic neuronal activity associated with motility in dog small intestine.

Intestinal motor activity associated with acetylcholine (ACh) release was assessed in the small intestine of anesthetized dogs by simultaneous measurement of motor activity and local ACh concentrations within the intestinal wall with in vivo microdialysis. Basal concentration of ACh measured in the dialysate was 1.12 +/- 0.08 pmol/15 min (n = 10), a value that remained constant until 3 h after perfusion. Intra-arterial infusion of tetrodotoxin reduced dialysate ACh concentration, while the motor activity accelerated at the early phase after infusion of tetrodotoxin and then decreased, thereby suggesting that the motor activity is regulated by not only excitatory cholinergic neurons, but also inhibitory neurons. Intraarterial infusion of atropine increased dialysate ACh concentration but reduced motor activity, thereby indicating that the cholinergic neurons are tonically active and the muscarinic autoreceptors operate to inhibit the ACh release. Intraarterial infusion of norepinephrine reduced, but yohimbine increased both motor activity and dialysate ACh concentration, thereby indicating that the adrenergic neurons regulate the motor activity due to control of cholinergic neuronal activity. This in vivo microdialysis method demonstrated in the whole body of animals that the activity of cholinergic neurons was physiologically regulated by itself and adrenergic neurons.

Effects of a domain peptide of the ryanodine receptor on Ca2+ release in skinned skeletal muscle fibers.

Mutations in the central domain of the skeletal muscle ryanodine receptor (RyR) cause malignant hyperthermia (MH). A synthetic peptide (DP4) in this domain (Leu-2442-Pro-2477) produces enhanced ryanodine binding and sensitized Ca2+ release in isolated sarcoplasmic reticulum, similar to the properties in MH, possibly because the peptide disrupts the normal interdomain interactions that stabilize the closed state of the RyR (Yamamoto T, El-Hayek R, and Ikemoto N. J Biol Chem 275: 11618-11625, 2000). Here, DP4 was applied to mechanically skinned fibers of rat muscle that had the normal excitation-contraction coupling mechanism still functional to determine whether muscle fiber responsiveness was enhanced. DP4 (100 microM) substantially potentiated the Ca2+ release and force response to caffeine (8 mM) and to low [Mg2+] (0.2 mM) in every fiber examined, with no significant effect on the properties of the contractile apparatus. DP4 also potentiated the response to submaximal depolarization of the transverse tubular system by ionic substitution. Importantly, DP4 did not significantly alter the size of the twitch response elicited by action potential stimulation. These results support the proposal that DP4 causes an MH-like aberration in RyR function and are consistent with the voltage sensor triggering Ca2+ release by destabilizing the closed state of the RyRs.

In vivo assessment of acetylcholine-releasing function at cardiac vagal nerve terminals.

We examined whether the ACh concentration measured by cardiac microdialysis provided information on left ventricular ACh levels under a variety of vagal stimulatory and modulatory conditions in anesthetized cats. Local administration of KCl (n = 5) and ouabain (n = 7) significantly increased the ACh concentration in the dialysate to 4.3 +/- 0.8 and 7.3 +/- 1.3 nmol/l, respectively, from the baseline value of 0.6 +/- 0.5 nmol/l. Intravenous administration of phenylbiguanide (n = 5) and phenylephrine (n = 6) significantly increased the ACh concentration to 5.4 +/- 0.9 and 6.0 +/- 1.5 nmol/l, respectively, suggesting that the Bezold-Jarisch and arterial baroreceptor reflexes affected myocardial ACh levels. Modulation of vagal nerve terminal function by local administration of tetrodotoxin (n = 6), hemicholinium-3 (n = 6), and vesamicol (n = 5) significantly suppressed the electrical stimulation-induced ACh release from 20.4 +/- 3.9 to 0.6 +/- 0.1, 7.2 +/- 1.9, and 2.7 +/- 0.6 nmol/l, respectively. Increasing the heart rate from 120 to 200 beats/min significantly reduced the myocardial ACh levels during electrical vagal stimulation, suggesting a heart rate-dependent washout of ACh. We conclude that ACh concentration measured by cardiac microdialysis provides information regarding ACh release and disposition under a variety of pathophysiological conditions in vivo.

In vivo microdialysis assessment of nerve-stimulated contractions associated with increased acetylcholine release in the dog intestine.

Intestinal contractility and release of endogenous acetylcholine (ACh) were measured simultaneously in vivo in the small intestine of the anesthetized dog. Electrical stimulation of nerves in the intestinal seromuscular layers caused contractions and increased concentrations of ACh in the dialysate, which were abolished by infusion of tetrodotoxin into the intestinal marginal artery at 75 nmol/ml. Intraarterial administration of atropine at 150 nmol/ml abolished the stimulated contractions, without significant effects on increases in concentrations of dialysate ACh. Thus, the nerve-stimulated contractions were found in vivo to be associated with a local increase in ACh release from the intestinal cholinergic neurons.

Altered sodium current response to intracellular fatty acids in halothane-hypersensitive skeletal muscle.

Biopsies of human skeletal muscle were analyzed by an in vitro contracture test (IVCT) for responsiveness to a halothane challenge: noncontracting (nonresponsive; IVCT-) and contracting (IVCT+). A muscle biopsy that is IVCT+ indicates potential malignant hyperthermia (MH) susceptibility. Primary cultures were grown from portions of the skeletal muscle biopsies, and voltage-activated currents were measured by whole cell recording in the presence or absence of 2-5 microM intracellular arachidonic or oleic acids. In untreated IVCT- cells, Na+ currents were predominantly tetrodotoxin (TTX) insensitive, indicating that most of the current was carried through the embryonic SkM2 isoform of the Na+ channel. Inclusion of fatty acids in the recording pipette of IVCT- cells produced an increase in voltage-activated Na+ currents during 20 min of recording. Approximately 70% of currents in fatty acid-treated cells were TTX sensitive, indicating activation of the adult SkM1 isoform of the Na+ channel. In contrast to IVCT- cells, IVCT+ cells expressed Na+ currents that were predominantly TTX sensitive even in the absence of added fatty acid, thus showing a relatively large baseline functional expression of SkM1 channels. Addition of fatty acids to the recording pipette produced little further change in the magnitude or TTX sensitivity of the whole cell currents in IVCT+ cells, suggesting altered functional regulation of Na+ channels in MH muscle.

Augmentation of prefrontal cortical monoaminergic activity inhibits dopamine release in the caudate nucleus: an in vivo neurochemical assessment in the rhesus monkey.

Prefrontal cortical modulation of caudate nucleus dopamine release was investigated in the rhesus monkey using the in vivo microdialysis technique. Reliable and stable basal caudate nucleus dopamine levels were quickly attained within hours following insertion of the dialysis probes. High-potassium (60 mM) or tetrodotoxin (10 microM) infusions significantly altered caudate nucleus dopamine levels in the dialysate indicating that measured dopamine levels reflected impulse-dependent release from the presynaptic pool. Pharmacological augmentation of monoaminergic transmission in the sulcus principalis region of the prefrontal cortex resulted in significant alterations in caudate nucleus dopamine levels. Increase of monoaminergic activity by infusion of either D-amphetamine (100 microM) or cocaine hydrochloride (100 microM) resulted in a gradual and prolonged decrease in caudate nucleus dopamine levels. Similar decreases were noticed in caudate nucleus dopamine metabolite levels. The present results indicate that in non-human primates modulation of dorsolateral prefrontal cortical monoaminergic transmission results in alterations in dopamine levels in subcortical structures. This observation may have clinical implications for therapeutic management of certain neuropsychiatric disorders, particularly schizophrenia.