Paraventricular noradrenergic systems participate in angiotensin II-induced drinking

The present study was designed to examine the role of noradrenergic systems in the hypothalamic paraventricular nucleus (PVN) in the drinking response induced by microinjection of angiotensin II (ANG II) into the subfornical organ (SFO) in the awake rat. Intracerebral microdialysis techniques were utilized to quantify the extracellular concentration of noradrenaline (NA) in the region of the PVN. Injections of ANG II (10⁻⁶ M, 0.2 μl) into the SFO significantly increased NA release in the PVN area. The increase in the NA concentration caused by the ANG II injection was significantly attenuated by water ingestion. In urethane-anesthetized rats, injections of ANG II into the SFO elicited an elevation in mean arterial pressure (MAP). On the other hand, intravenous injections of the -agonist metaraminol (5 μg) slightly decreased the release of NA in the PVN area that accompanied an elevation in MAP. These results show that the noradrenergic system in the PVN area may be involved in the dipsogenic response induced by ANG II acting at the SFO.     

Nutritive blood flow improves interstitial glucose and lactate exchange in perfused rat hindlimb

Microdialysis was used to assess the interstitial concentrations of glucose and lactate in the constant-flow-perfused rat hindlimb under varying levels of nutritive flow controlled by vasoconstrictors. Increased nutritive flow was achieved by norepinephrine (NE) or angiotensin II (ANG II) and decreased nutritive flow by serotonin (5-HT). NE and ANG II increased oxygen and glucose uptake as well as hindlimb lactate release by 50%. 5-HT decreased oxygen uptake by 15% but had no significant effect on glucose uptake or hindlimb lactate release. Microdialysis recovery of glucose and lactate was significantly elevated by NE and ANG II and decreased by 5-HT. The calculated interstitial concentration of glucose was increased by NE and ANG II but decreased by 5-HT. The interstitial concentration of lactate was decreased by NE and ANG II but increased by 5-HT. In all cases, nitroprusside reversed the effects of the vasoconstrictors. These data indicate that increased nutritive blood flow enhances the exchange of glucose and lactate by improving the supply of glucose to and the removal of lactate from the interstitium.     

Central actions of angiotensin in cardiovascular control: multiple roles for a single peptide.

Angiotensin II (ANG II) acts peripherally as a hormone, with actions on the vasculature, adrenals, and kidney. In addition, certain actions of ANG II in the central nervous system are directed toward cardiovascular control and fluid volume homeostasis. Dense binding sites for ANG II are found at circumventricular organs, which apparently have the ability to relay information to cardiovascular centers via neural circuitry. Microinjection of ANG II into the subfornical organ (SFO) or area postrema (AP) produces site-specific increases in blood pressure. In addition, electrophysiological studies demonstrate profound effects of ANG II, acting at the SFO, on activity of neurohypophysial neurons and release of oxytocin and vasopressin, which can be antagonized by ANG II blockers or attenuated by SFO lesions. Evidence from microinjection, electrophysiological, and lesion studies indicate a complex interaction between central sites involved in mechanisms of cardiovascular control: the SFO, AP, organum vasculosum of the lamina terminalis, and paraventricular and supraoptic nuclei of the hypothalamus. Not only is ANG II a humoral messenger in this central scenario, but evidence suggests it acts as a neurotransmitter or neuroendocrine substance within specific CNS pathways, suggesting multiple roles for this peptide in central cardiovascular control.     

Decreased monoamine release in the median preoptic area following ventricular treatment with the angiotensin II antagonist saralasin in normotensive and spontaneously hypertensive rats

Previous findings have shown that some of the neurons in the median preoptic nucleus (MnPO) receive both catecholaminergic inputs from the brainstem and angiotensinergic inputs from the subfornical organ (SFO), and that alterations in the function of the brain renin-ANG system are implicated in hypertension, especially in spontaneous hypertensive rats (SHR). In an attempt to clarify the action of these inputs on MnPO neurons and to find the difference in the action between normotensive Wistar-Kyoto (WKY) rats and SHR, we used microdialysis to investigate the effects of injections of saralasin (Sar), an angiotensin II (ANG II) antagonist, into the third ventricle (3V) on monoamine release in the MnPO area of awake WKY and SHR. The content of noradrenaline (NA) in the MnPO area was significantly higher in SHR. No significant differences were observed between WKY and SHR in the concentrations of dopamine (DA) and of its two metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). In both WKY and SHR, Sar (Sar, 5 microg in 1 microl, three injections at 2-h intervals) injected into the 3V significantly decreased the extracellular concentrations of NA, DOPAC and HVA in the MnPO area. The decreases were much greater in SHR than in WKY rats. Similar injections of saline vehicle had no significant effect on the extracellular levels of NA, DA and the metabolites. These results suggest that central angiotensinergic circuits may serve to increase NA and DA release in the MnPO area, and support that a disorder in the ANG system may contribute, in part, to the elevated blood pressure of SHR.     

Circumventricular organs and ANG II-induced salt appetite: blood pressure and connectivity

A lesion of the subfornical organ (SFO) may reduce sodium depletion-induced salt appetite, which is largely dependent on ANG II, and yet ANG II infusions directly into SFO do not provoke salt appetite. Two experiments were designed to address this apparent contradiction. In experiment 1 sustained infusions of ANG II into SFO did not produce a sustained elevation of blood pressure, and neither a reduction of blood pressure alone with minoxidil and captopril nor a reduction of both blood pressure and volume with furosemide and captopril enhanced salt appetite. Infusions of ANG II in the organum vasculosum laminae terminalis (OVLT) did evoke salt appetite without raising blood pressure. In experiment 2 knife cuts of the afferent and efferent fibers of the rostroventral pole of the SFO abolished water intake during an infusion of ANG II into the femoral vein but failed to reduce salt appetite during an infusion of ANG II into the OVLT. We conclude that 1) hypertension does not account for the failure of infusions of ANG II in the SFO to generate salt appetite and 2) the OVLT does not depend on its connectivity with the SFO to generate salt appetite during ANG II infusions.     

Contribution of the subfornical organ to angiotensin II-induced hypertension

Previous studies clearly demonstrated acute actions of angiotensin II (ANG II) at one of the central circumventricular organs, the subfornical organ (SFO), but studies demonstrating a role for the SFO in the chronic actions of ANG II remain uncertain. The purpose of this study was to examine the role of the SFO in the chronic hypertensive phase of ANG II-induced hypertension. We hypothesized that the SFO is necessary for the full hypertensive response observed during the chronic phase of ANG II-induced hypertension. To test this hypothesis, male Sprague-Dawley rats were subjected to sham operation (sham rats) or electrolytic lesion of the SFO (SFOx rats). After 1 wk, the rats were instrumented with venous catheters and radiotelemetric transducers for intravenous administration of ANG II and measurement of blood pressure and heart rate, respectively. Rats were then allowed 1 wk for recovery. After 3 days of saline control infusion (7 ml of 0.9% NaCl/day), sham and SFOx rats were infused with ANG II at 10 ng.kg(-1).min(-1) i.v. for 10 consecutive days and then allowed to recover for 3 days. A 0.4% NaCl diet and distilled water were provided ad libitum. At day 5 of ANG II infusion, mean arterial pressure increased 11.7 +/- 3.0 mmHg in sham rats (n = 9) but increased only 3.7 +/- 1.4 mmHg in SFOx rats (n = 9). This trend continued through day 10 of ANG II treatment. These results support the hypothesis that the SFO is necessary for the full hypertensive response to chronic ANG II administration.     

Subfornical organ differentially modulates baroreflex function in normotensive and two-kidney, one-clip hypertensive rats

During activation of the renin-angiotensin system, hindbrain circumventricular organs such as the area postrema have been implicated in modulating the arterial baroreflex. This study was undertaken to test the hypothesis that the subfornical organ (SFO), a forebrain circumventricular structure, may also modulate the baroreflex. Studies were performed in rats with two-kidney, one-clip (2K,1C) hypertension as a model of endogenously activated renin-angiotensin system. Baroreflex function was ascertained during ramp infusions of phenylephrine and nitroprusside in conscious sham-clipped and 5-wk 2K,1C rats with either a sham or electrolytically lesioned SFO. Lesioning significantly decreased mean arterial pressure in 2K,1C rats from 158 +/- 7 to 131 +/- 4 mmHg but not in sham-clipped rats. SFO-lesioned, sham-clipped rats had a significantly higher upper plateau and range of the renal sympathetic nerve activity-mean arterial pressure relationship compared with sham-clipped rats with SFO ablation. In contrast, lesioning the SFO in 2K,1C rats significantly decreased both the upper plateau and range of the baroreflex control of renal sympathetic nerve activity, but only the range of the baroreflex response of heart rate decreased. Thus, during unloading of the baroreceptors, the SFO differentially modulates the baroreflex responses in sham-clipped vs. 2K,1C rats. Since lesioning the SFO did not influence plasma angiotensin II (ANG II), the effects of the SFO lesion are not caused by changes in circulating levels of ANG II. These findings support a pivotal role for the SFO in the sympathoexcitation observed in renovascular hypertension and in baroreflex regulation of sympathetic activity in both normal and hypertensive states.     

Changes in plasma and brain tryptophan and brain serotonin and 5-hydroxyindoleacetic acid after footshock stress

Brain concentrations of tryptophan, serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) and plasma amino acids were measured after 15 or 30 minutes of intermittent footshock. Footshock treatment significantly decreased the content of 5-HT in prefrontal cortex and hypothalamus, but not brainstem at 15 min, but the decreases were reversed by 30 min. 5-HIAA, the major catabolite of 5-HT, increased in prefrontal cortex after 15 min, and in prefrontal cortex and hypothalamus after 30 min footshock. 5-HIAA:5-HT ratios were increased at both timepoints in all three brain regions. Concomitant changes in the ratios of 3,4-dihydroxyphenylacetic acid (DOPAC) to dopamine and 3-methoxy,-4-hydroxyphenylethyleneglycol (MHPG) to norepinephrine were also observed. Brain concentrations of tryptophan increased progressively during the footshock in all three brain regions. Plasma concentrations of both tryptophan and tyrosine were also significantly increased, while those of histidine and lysine were decreased. It is possible that the stress-related changes in 5-HT metabolism are due to increased plasma tryptophan, in turn causing increased brain tryptophan and 5-HT synthesis. However, the transient decreases in 5-HT suggest a footshock-induced increase of 5-HT release, depleting existing stores of 5-HT, that are replenished by the increased systemic availability of tryptophan.     

Neocortical Dialysate Monoamines of Rats After Acute, Subacute, and Chronic Liver Shunt

Abstract: Intracerebral microdialysis was applied to monitor the neocortical extracellular levels of the aromatic amino acids phenylalanine, tyrosine, and tryptophan, the neurotransmitters dopamine (DA), noradrenaline (NA), and serotonin (5-HT), and the metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and 5-hydroxyindole-3-acetic acid (5-HIAA) in rats with various forms of experimental hepatic encephalopathy (HE). The extracellular aromatic amino acid levels were clearly increased in acute, subacute, and chronic HE. No changes compared with controls in the neocortical DA release could be detected in the three experimental HE rat models investigated. The NA release showed a significant increase only in the subacute HE group. These data suggest that HE may not be associated with any major reduction of neocortical DA or NA release as previously suggested. In acute and subacute HE, decreased extracellular DOPAC but elevated 5-HIAA concentrations were seen. In chronic HE, elevations of both DOPAC and 5-HIAA were observed. Neocortical 5-HT release did not change in subacute and chronic HE, whereas it decreased in acute HE compared with control values. Significant increase in extracellular concentrations of 5-HIAA and of the 5-HIAA/5-HT ratio in the present study are in agreement with previously reported increases in 5-HT turnover in experimental HE. However, a substantially increased 5-HT turnover in experimental HE does not appear to be related to an increase in neuronal neocortical 5-HT release.     

Median preoptic nucleus and subfornical organ drive renal sympathetic nerve activity via a glutamatergic mechanism within the paraventricular nucleus

The paraventricular nucleus (PVN) of the hypothalamus is involved in the neural control of sympathetic drive, but the precise mechanism(s) that influences the PVN is not known. The activation of the PVN may be influenced by input from higher forebrain areas, such as the median preoptic nucleus (MnPO) and the subfornical organ (SFO). We hypothesized that activation of the MnPO or SFO would drive the PVN through a glutamatergic pathway. Neuroanatomical connections were confirmed by the recovery of a retrograde tracer in the MnPO and SFO that was injected bilaterally into the PVN in rats. Microinjection of 200 pmol of N-methyl-d-aspartate (NMDA) or bicuculline-induced activation of the MnPO and increased renal sympathetic activity (RSNA), mean arterial pressure, and heart rate in anesthetized rats. These responses were attenuated by prior microinjection of a glutamate receptor blocker AP5 (4 nmol) into the PVN (NMDA - ΔRSNA 72 ± 8% vs. 5 ± 1%; P < 0.05). Using single-unit extracellular recording, we examined the effect of NMDA microinjection (200 pmol) into the MnPO on the firing activity of PVN neurons. Of the 11 active neurons in the PVN, 6 neurons were excited by 95 ± 17% (P < 0.05), 1 was inhibited by 57%, and 4 did not respond. The increased RSNA after activation of the SFO by ANG II (1 nmol) or bicuculline (200 pmol) was also reduced by AP5 in the PVN (for ANG II - ΔRSNA 46 ± 7% vs. 17 ± 4%; P < 0.05). Prior microinjection of ANG II type 1 receptor blocker losartan (4 nmol) into the PVN did not change the response to ANG II or bicuculline microinjection into the SFO. The results from this study demonstrate that the sympathoexcitation mediated by a glutamatergic mechanism in the PVN is partially driven by the activation of the MnPO or SFO.     

Imidazoline receptors of the paraventricular nucleus on the pressor response induced by stimulation of the subfornical organ

In the present experiments we investigated a possible involvement of imidazoline receptors of the paraventricular nucleus (PVN) of the hypothalamus on the pressor effects of the angiotensin II (ANG II) injected into the subfornical organ (SFO), in male Holtzman rats (250–300 g) with a cannula implanted into the third ventricle (3rdV), PVN and SFO. At first we tested the participation of α₂ and imidazoline agonist and antagonist compounds on the pressor effect of ANG II injected into the 3rdV. Based on the results we may conclude that clonidine associated with rilmenidine was able to block the hypertensive response to ANG II. The ANG II (20 pmol) injected into SFO induced a robust increase in blood pressure (37 ± 2 mmHg). Isotonic saline (0.15 M) NaCl did not produce any change in blood pressure (5 ± 2 mmHg). The injection of rilmenidine (30 μg/kg/1 μL), an imidazoline agonist agent injected into PVN before ANG II injection into SFO, blocked the pressor effect of ANG II (5 ± 2 mmHg). Also, the injection of idazoxan (60 μg/kg/μL) before rilmenidine blocked the inhibitory effect of rilmenidine on blood pressure (39 ± 4 mmHg). The injection of clonidine (20 nmol/μL) prior to ANG II into the 3rdV produced a decreased in arterial blood pressure (37 ± 2 mmHg) to (15 ± 4 mmHg). The injection of yohimbine (80 nmol/μL) prior to clonidine blocked the effect of clonidine on the effect of ANG II (27 ± 2 mmHg). The injection of rilmenidine prior to ANG II also induced a decrease in arterial blood pressure (10 ± 3 mmHg). The injection of idazoxan prior to rilmenidine also blocked the inhibitory effect of rilmenidine (24 ± 3 mmHg). In summary, the present study demonstrated that rilmenidine decreases the hypertensive effect of ANG II, with more potency than clonidine, even when injected into 3rdV or PVN. This study established that the PVN interacts with SFO by imidazoline receptors in order to control the arterial blood pressure.     

In vivo release of endogenous dopamine, 5-hydroxytryptamine and some of their metabolites from rat caudate nucleus by phenylethylamine

The in vivo release of endogenous 3,4-dihydroxyphenylethylamine (DA) and its metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA) and 3-methoxytyramine (3-MT), and of 5-hydroxytryptamine (5-HT) and its metabolite, 5-hydroxyindoleacetic acid (5-HIAA), has been measured in the caudate nucleus of the anesthetized rat. A push-pull cannula was implanted into the brain, and the tissue perfused with artificial CSF or artificial CSF containing 5×10^–4 M phenylethylamine. The perfusate was collected and analyzed for DA, 5-HT and their metabolites by high performance liquid chromatography with electrochemical detection (HPLC-ECD). DA was released by phenylethylamine at rates significantly greater than its basal rate. 3-MT and 5-HT were undetectable in perfusates collected under basal conditions, but could be detected readlly during phenylethylamine stimulation. DOPAC, HVA and 5-HIAA concentrations were not significantly affected by phenylethylamine. The results suggest (1) that phenylethylamine may exert its behavioural effects through increased release of both DA and 5-HT, and (2) that in vivo measurements of the acid metabolites alone may not be indicative of the release of the amines.Special Issue Dedicated to Dr. Abel Lajtha.     

Exogenous Tryptophan Increases Synthesis, Storage, and Intraneuronal Metabolism of 5-Hydroxytryptamine in the Rat Hypothalamus

The effects of tryptophan administration on neurochemical estimates of synthesis [5-hydroxytryptophan (5-HTP) accumulation following administration of a decarboxylase inhibitor], storage [5-hydroxytryptamine (5-HT) concentrations], and metabolism [5-hydroxyindoleacetic acid (5-HIAA) concentrations] of 5-HT in selected regions of the hypothalamus were determined using HPLC coupled to an electrochemical detector. Tryptophan methyl ester HCl (30-300 mg/kg i.p.) produced a dose-dependent increase in the rate of 5-HTP accumulation throughout the hypothalamus but had no effect on the rate of accumulation of 3,4-dihydroxyphenylalanine. Peak 5-HTP levels were attained by 30 min following administration of tryptophan (100 mg/kg i.p.) and were maintained for an additional 60 min. Tryptophan also produced concomitant dose-dependent increases in 5-HT and 5-HIAA concentrations in these same regions without changes in the 5-HIAA/5-HT ratio. These results indicate that exogenous tryptophan administration selectively increases the synthesis, storage, and metabolism of 5-HT in the hypothalamus without altering the synthesis of catecholamines. Inhibition of 5-HT uptake with chlorimipramine or fluoxetine produced modest (10-40%) reductions in 5-HIAA concentrations throughout the hypothalamus, revealing that only a minor portion of 5-HIAA is derived from released and recaptured 5-HT, whereas the major portion of this metabolite reflects intraneuronal metabolism of unreleased 5-HT. In both chlorimipramine- and fluoxetine-treated rats, 5-HIAA concentrations were significantly increased by tryptophan administration, indicating that the increase in synthesis of 5-HT following precursor loading is accompanied by an increase in the intraneuronal metabolism of 5-HT.     

Effect of NMDA-Induced Lesion of the Subfornical Organ on the Angiotensin II Binding Sites Density and Acetylcholinesterase or NADPH-Diphorase Activities in the Lamina Terminalis of the Rat Brain

1. Neural angiotensinergic circuitry located in the lamina terminalis has been proposed to be involved in blood pressure regulation and fluid homeostasis.2. ANG II binding sites have been described to be localized throughout the lamina terminalis including the subfornical organ (SFO), the median preoptic nucleus (MnPO), and the organum vasculosum lamina terminalis (OVLT).3. The present experiment was designed to investigate the ANG II binding sites localization in the lamina terminalis. For this purpose, we have compared the ANG II binding sites, acetylcholinesterase, and NADPH-diaphorase distributions throughout the lamina terminalis. Additionally, we have studied the effect ofthe preferential lesion of SFO neuronal cell bodies by local injection of NMDA on the ANG II binding sites density in different areas of the lamina terminalis.4. Male Wistar rats were anesthetized, immobilized in a stereotaxic apparatus, and 500 nl of saline or 250 nmol NMDA was injected into the SFO.5. Animals were sacrificed 1 week later, the brain was removed, frozen, and sagittal 16 m slices were cut in a cryostat. Alternate brain slices were incubated with [¹²⁵I]-Sar¹-ANG II for receptor autoradiography or histochemically stained for visualization of acetylcholinesterase and NADPH-diaphorase activities. Binding capacity was determined by computerized quantitative densitometry of autoradiograms. The intensity of histochemical reactions was measured as relative units obtained by computerized densitometry processing of the brain slices stained for either activity.6. Acetylcholinesterase staining was mainly located in the SFO, with faint staining reaction in other areas of the lamina terminalis. NADPH-diaphorase staining was homogeneously distributed throughout the lamina terminalis. A significant positive correlation was observed between acetylcholinesterase and NADPH-diaphorase stainings in the SFO of control and NMDA-lesioned rats.ANG II binding sites were localized throughout the lamina terminalis. A significant positive correlation was observed between the density of ANG II binding sites and the intensity of acetylcholinesterase or NADPH-diaphorase staining in the SFO of control and NMDA-lesioned rats.8. The distribution of the NADPH-diaphorase staining was found to closely match the distribution of the ANG II binding sites in the lamina terminalis.9. Neuronal lesion of the SFO caused significant reductions in the density of ANG II biding sites in the SFO (–68%) and the MnPO (–48%). No changes were observed either in the OVLT or outside the lamina terminalis in the superior colliculus.10. The present results indicate the following: first, the presence ofhigh levels of acetylcholinesterase staining in the SFO and of NADPH-diaphorasethroughout the lamina terminalis; second, that ANG II binding sites in the SFO and possibly in the MnPO are localized in neuronal cell bodies; third, that SFOlesion did not affect the expression of ANG II binding sites in the OVLT, thus suggesting that these binding sites correspond to different angiotensinergic system; and finally, the existence of a striking correlation between the distribution of the ANG II binding sites and NADPH-diaphorase throughout the lamina terminalis, thus suggesting a interrelation between angiotensinergic and nitrergicsystems in the lamina terminalis.     

Evidence for involvement of 5-hydroxytryptamine(1B) autoreceptors in the enhancement of serotonin turnover in the mouse brain following repeated treatment with fluoxetine

The effect of repeated treatment with the selective serotonin reuptake inhibitor fluoxetine on synthesis and turnover of 5-hydroxytryptamine (5-HT) was studied in the mouse brain in vivo. The concentration of 5-hydroxytryptophan (5-HTP), 5-hydroxyindoleacetic acid (5-HIAA) and 5-HT was measured in hypothalamus, hippocampus and frontal cortex after inhibition of the aromatic amino acid decarboxylase activity with m-hydroxybenzylhydrazine (NSD 1015). Fluoxetine 6.9 mg/kg s.c. was injected once daily for three weeks. Three days after the final daily injection of fluoxetine 5-HT synthesis (5-HTP accumulation) and turnover (5-HIAA/5-HT ratio) were significantly enhanced compared with saline-treated mice. The 5-HIAA/5-HT ratio was already significantly elevated after 3 days of fluoxetine treatment and continued to increase during treatment for 2-3 weeks. The increase in 5-HIAA/5-HT ratio was considerably larger (150-200% of controls) than the increase in 5-HTP accumulation (110-120%), which reached significance only after 3 weeks of treatment. The increase in 5-HT synthesis may be secondary to that of the turnover. The 5-HIAA/5-HT ratio returned to control values after a 14 days washout period. Simultaneous treatment with the long-acting 5-HT(1B)-receptor antagonist, SB 224289 for 14 days counteracted the fluoxetine-induced increase in 5-HIAA/5-HT ratio that indicates involvement of 5-HT(1B) autoreceptors in the development of this increase. It is proposed that the fluoxetine-induced enhancement of 5-HT turnover was evoked by the long-lasting stimulation of 5-HT(1B) autoreceptors that resulted in an intraneuronal compensatory adaptation of the basal 5-HT release.     

Calcium-Dependent, Tetrodotoxin-Sensitive Stimulation of Cortical Serotonin Release After a Tryptophan Load

The effect of intraperitoneal administration of tryptophan (50, 100, or 200 mg/kg) on extracellular concentrations of tryptophan, serotonin (5-hydroxytryptamine, 5-HT), and 5-hydroxyindoleacetic acid (5-HIAA) was studied in the cortex of freely moving rats by transcerebral dialysis. Rats were implanted with dialysis probes in the frontal cortex, and experiments were performed 24 h later. Tryptophan, 5-HT, and 5-HIAA were quantified in 20-min samples of dialysate by HPLC with electrochemical detection after separation on reverse-phase columns. Tryptophan administration resulted in a significant increase of tryptophan, 5-HT, and 5-HIAA levels in dialysates. The maximal increase of 5-HT and 5-HIAA output was approximately 150% over basal values. Perfusion with Ringer's solution containing tetrodotoxin (1 microM) reduced 5-HT output by 90% and prevented the increase of 5-HT and 5-HIAA content after 100 mg/kg of tryptophan. Similar results were obtained after perfusion with Ringer's solution without Ca2+. The results indicate that a tryptophan load stimulates the physiological release of 5-HT.     

Evidence for GABA control of serotonin metabolism in the rat suprachiasmatic area

Changes in endogenous serotonin (5-HT) metabolism after in vivo stimulation of GABAergic transmission were investigated in the rat suprachiasmatic area (SCA). Activation of GABA transmission was performed by systemic administration of either amino-oxyacetic acid: AOAA, a GABA-transaminase inhibitor or RS baclofen, a GABA B agonist. After drugs administration, the amounts of endogenous 5-HT and 5-HIAA were measured. The release and synthesis of 5-HT were investigated in vitro, using a static incubation of tissue fragments. AOAA or RS baclofen induced an increase in endogenous 5-HT content but did not affect 5-hydroxyindole-acetic acid (5-HIAA). Both drugs induced an increase in the release and synthesis of 5-HT. Detailed study of the effects of AOAA over time on 5-HT metabolism showed that the increase in 5-HT release preceded the increase in amine synthesis. These results suggest that the in vivo stimulation of GABA transmission induces an increase in metabolic activity of the 5-HT neuronal system in the SCA. This effect may likely be mediated via activation of GABA B receptors.     

Peripheral interleukin-6 administration increases extracellular concentrations of serotonin and the evoked release of serotonin in the rat striatum

Previous studies have indicated that peripheral administration of interleukin-6 (IL-6) increases brain concentrations of tryptophan and 5-hydroxyindoleacetic acid (5-HIAA), the major catabolite of serotonin (5-HT). To determine whether these changes were related to increased synaptic release of 5-HT, we studied the responses to peripheral administration of IL-6 by in vivo microdialysis and in vivo amperometry. Intraperitoneal injection of recombinant IL-6 resulted in an elevation of microdialysate concentrations of 5-HT in the rat striatum. Also, amperometric measurements indicated that i.p. IL-6 enhanced the 5-HT-like signal obtained from the striatum following electrical stimulation of the dorsal raphe nucleus. These results indicate that the increases in brain concentrations of 5-HIAA observed in earlier studies indeed reflect increased synaptic release of 5-HT.     

Cerebrospinal fluid monoamine and metabolite concentrations and aggression in rats

In humans and other primates low cerebrospinal fluid (CSF) levels of the major serotonin (5-HT) metabolite 5-hydroxyindoleacetic acid (5-HIAA) have been correlated to high aggressiveness. This finding forms the basis of the 5-HT deficiency hypothesis of aggression. Surprisingly, this correlation has not been confirmed in rodents so far, while manipulation studies aimed to investigate the link between 5-HT and aggressive behaviour are mostly carried out in rodents. In this study the relation between aggression and CSF monoamine and metabolite concentrations was investigated in male Wildtype Groningen rats. In sharp contrast to the hypothesis and our expectation, a clear positive correlation was found between the individual level of trait-like aggressiveness and CSF concentrations of 5-HT, 5-HIAA, norepinephrine (NE), dopamine (DA), and 3,4-dihydroxyphenylacetic acid (DOPAC). Shortly after the acute display of aggressive behaviour (as a state-like phenomenon), decreased 5-HT levels and an increase in 5-HIAA/5-HT ratio and NE concentrations were found. Surprisingly, pharmacological challenges known to influence 5-HT transmission and aggressive behaviour did not affect CSF 5-HT and 5-HIAA concentrations, only the NE level was increased. Lesioning 5-HT terminals by 5,7-dihydroxytryptamine (5,7-DHT) administration caused a decrease in CSF 5-HT and 5-HIAA, but without affecting aggressive behaviour. The observed positive correlation between CSF 5-HIAA and trait aggressiveness makes it questionable whether a direct extrapolation of neurobiological mechanisms of aggression between species is justified. Interpretation of CSF metabolite levels in terms of activity of neural substrates requires a far more detailed knowledge of the dynamics and kinetics of a neurotransmitter after its release.     

Simultaneous Effects of p-Chloroamphetamine, d-Fenfluramine, and Reserpine on Free and Stored 5-Hydroxytryptamine in Brain and Blood

The effects of acute treatment with p-chloramphetamine, d-fenfluramine, and reserpine on intracellular (brain tissue and whole blood) and extracellular (CSF and platelet-free plasma) compartments of 5-hydroxytryptamine (5-HT) in the brain and blood of the same rats have been examined. These treatments affected 5-HT in brain tissue and whole blood similarly (r = 0.823). Reserpine significantly reduced both intracellular pools at 2 and 24 h. p-Chloroamphetamine and d-fenfluramine were more effective on brain tissue 5-HT. The concentration of 5-HT in CSF was significantly increased by all treatments. p-Chloroamphetamine induced a dramatic 70-fold increase of CSF 5-HT, paralleling a 42% decrease in brain tissue. d-Fenfluramine significantly increased CSF 5-HT to 212% of controls and reduced whole brain 5-HT (-23%). The effects of p-chloroamphetamine and d-fenfluramine on 5-HIAA in brain, CSF, and plasma were nonsignificant. Individual values of 5-hydroxyindoleacetic acid (5-HIAA) in CSF and brain were highly correlated (r = 0.855), indicating that CSF 5-HIAA reflects well the concentration of 5-HIAA in brain tissue. Yet the intra- and extracellular concentrations of 5-HIAA were unrelated to the 5-HT changes. This indicates that CSF 5-HIAA does not reflect the active (extracellular) compartment of 5-HT in brain.