Angiotensin II-induced noradrenaline release from anterior hypothalamus in conscious rats: a brain microdialysis study.

Interaction with aminergic transmitter substances has been implicated in the central actions of angiotensin II (ANG II). We used the novel technique of brain microdialysis in conscious rats to investigate whether ANG II influences the release of endogenous catecholamines (CA) from the anterior hypothalamus (AH). Intracerebroventricular (i.c.v.) administration of ANG II (1 ng and 100 ng) increased mean arterial pressure. ANG II at 1 ng had no effect on the release of noradrenaline (NA) from the AH but ANG II at 100 ng significantly increased NA release. Intracerebroventricular pretreatment with the ANG II-receptor antagonist sarilesin (Sar1, Ile8-ANG II; 3 micrograms) prevented the ANG II-induced NA release. The release of the intraneuronal NA and dopamine (DA) metabolites 3,4-dihydroxyphenylethyl glycol (DOPEG) and 3,4-dihydroxyphenylacetic acid (DOPAC) from the AH was not altered by i.c.v. ANG II. Our results provide the first in vivo evidence for NA release from the hypothalamus induced by periventricular ANG II receptor stimulation. They support the hypothesis that hypothalamic noradrenergic mechanisms are involved in the central actions of this peptide.     

Somatosensory stimuli evoke norepinephrine release in the anterior ventromedial hypothalamus of sexually receptive female rats

We used in vivo brain microdialysis to determine the role of specific copulatory stimuli in mating-induced release of norepinephrine in the lateral ventromedial hypothalamus (VMH) of hormone-treated, sexually receptive female rats. Ovariectomized rats implanted with a unilateral guide cannula aimed at the ventrolateral VMH received systemic injections of estradiol benzoate daily for 2 days before and progesterone 4 h before the initiation of a 1-h behavioural test. Dialysis probes were lowered immediately after progesterone administration, and 20-min dialysis samples were collected until 1 h after the termination of behavioural testing. Norepinephrine content of dialysates was quantified by high performance liquid chromatography with electrochemical detection. During mating tests with male rats, dialysate levels of norepinephrine increased significantly over baseline in sexually receptive females with probe placements in the anterior but not posterior VMH. Norepinephrine levels were unchanged if rats were nonreceptive, even if males mounted vigorously and probes were located in the anterior VMH. Hormone-treated females that were placed on male-soiled bedding for 1 h showed no changes in dialysate levels of norepinephrine. Similarly, females in which vaginocervical stimulation was prevented by a vaginal mask failed to show increased levels of norepinephrine in dialysates collected from the anterior VMH, even if they displayed high levels of lordosis behaviour. Thus, the release of norepinephrine is not a result of executing the lordosis posture. The findings suggest that mating-induced increases in norepinephrine release in hormone-treated, sexually receptive rats are confined to the anterior VMH and that somatosensory rather than chemosensory stimuli evoke norepinephrine release. Moreover, experiments with vaginal masks indicate that vaginocervical stimulation is necessary for mating-evoked norepinephrine release in the anterior VMH.     

Activation of descending noradrenergic system by peripheral nerve stimulation

Noradrenaline (NA) release in the rat lumbar spinal cord (L3–4) in response to variable intensity, selective stimulation of large (A-beta), small myelinated (A-delta), and unmyelinated (C) afferent fibers was examined by in vivo microdialysis with high performance liquid chromatography and electrochemical detection. Application of 100 mM K⁺ solution via the dialysis probe increased NA in the dialysate. Thoracic segment transection rostral to the probe depressed the NA level. Transcutaneous stimulation of peripheral nerves had the following effects: 1) High intensity stimulation of afferent A-delta or C fibers increased spinal NA release, which was decreased by thoracic spinal cord transection. 2) Stimulation of afferent A-beta or A-delta fibers at low intensity did not affect the NA level. 3) High intensity stimulation of afferent A-beta fibers depressed NA release in half of the trials. Results indicate that many NA-containing nerve terminals that innervate the lumbar spinal cord originate from supraspinal structures. Somatic neural inputs from afferent C fibers and high-threshold A-delta, but not A-beta nor low-threshold A-delta fibers, activate the descending NA system and release the NA in the spinal cord. The descending NA system may participate in antinociception.     

Puberty acceleration in female mice induced with a partially purified male urine extract: effects on catecholamine release from the olfactory bulbs and hypothalamus

In the present experiment peri-pubertal female mice were treated with a partially purified puberty accelerating urine extract (PAUE). Mice treated with the PAUE showed an advance in the onset of puberty as indicated by significantly increased uterine weights. Treatment with the PAUE did not alter basal or potassium- (K+, 30 mM) stimulated release of catecholamines (dopamine or norepinephrine) from either anterior or posterior superfused olfactory bulb tissue fragments. There was, however, an overall significantly greater amount of basal and K(+)-stimulated release of NE from the posterior vs. the anterior olfactory bulb. Potassium-stimulated-, but not basal, release of catecholamines from the medial basal hypothalamus of PAUE-treated female mice were increased, with dopamine showing a statistically significant difference compared to water-treated females. These data demonstrate that treatment with the PAUE is a very effective means to accelerate the onset of puberty and results in accompanying increases in catecholaminergic activity, in particular dopamine, within the medial basal hypothalamus.     

Endotoxin administration stimulates cerebral catecholamine release in freely moving rats as assessed by microdialysis

In vivo microdialysis was used to measure changes in extracellular concentrations of catecholamines and indolamines in freely moving rats in response to administration of endotoxin (lipopolysaccharide, LPS). Dialysis probes were placed stereotaxically in either the medial hypothalamus or the medial prefrontal cortex. We used a repeated-measures design in which each rat received LPS or saline, and each subject was retested with the other treatment one week later. With the dialysis probes in the medial hypothalamus, intraperitoneal (ip) administration of LPS (5 μg) increased dialysate concentrations of norepinephrine (NE, 187%), dopamine (DA, 119%), and all their measured catabolites, except normetanephrine. Dialysate concentrations of NE and DA were elevated significantly in the fourth or fifth (20 min) collection period with a peak response at around 2 hr. They returned to baseline by about 4 hr. When the dialysis probes were placed in the medial prefrontal cortex, the same dose of LPS also elevated dialysate concentrations of NE and DA, but the increases were much smaller (ca. 20%). However, a dose of 100 μg LPS increased dialysate concentrations of NE and DA from the medial prefrontal cortex to an extent comparable to that of the 5 μg dose in the hypothalamus, and the response was more prolonged. Dialysate concentrations of serotonin could not be measured reliably, but those of its catabolite, 5-hydroxyindoleacetic acid (5-HIAA), were also elevated in both regions. The peak of 5-HIAA occurred at around 4 hr. Pretreatment of the rats with indomethacin (10 mg/kg ip) completely prevented the changes due to 100 μg LPS in the medial prefrontal cortex. These results support earlier neurochemical data suggesting that LPS stimulates the release of both DA and NE in the brain, and probably also release of serotonin. © 1995 Wiley-Liss, Inc.     

In vivo assessment of catechol O-methyltransferase activity in rabbit skeletal muscle

With the use of microdialysis technique in the anesthetized rabbit, we examined the catechol O-methyltransferase (COMT) activity at the skeletal muscle interstitium. We implanted a dialysis probe into the adductor muscle, and monitored dialysate catecholamines and their metabolites with chromatogram-electrochemical detection. Administration of COMT inhibitor (entacapone) decreased dialysate 3-methoxy 4-hydroxyphenylglycol (MHPG) levels. Local administration of dihydroxyphenylglycol induced increases in dialysate MHPG levels. These increases in dialysate MHPG levels were suppressed by the addition of entacapone. The concentration of MHPG in the skeletal muscle dialysate corresponded to the COMT activity in the skeletal muscle. Furthermore, local administration of norepinephrine or epinephrine increased normetanephrine or metanephrine levels in dialysate but not MHPG levels. Skeletal muscle microdialysis with local administration of catecholamine offers a new method for in vivo assessment of regional COMT activity.     

Central and systemic oxytocin release: A study of the paraventricular nucleus by in vivo microdialysis

The mechanisms controlling central and systemic oxytocin (OT) release were examined using in vivo microdialysis of the paraventricular (PVN) region. Dialysate and plasma samples were collected from conscious male rats and stimuli were administered via the dialysate fluid. Characterization studies showed that microdialysis was a viable technique for the study of peptide secretion in the conscious animal. OT was consistently detected in the PVN dialysate and a partially purified extract crossreacted in parallel fashion with the synthetic peptide. In vitro studies showed that peptide recovery was positively correlated with the pore size of the dialysis membrane and that there was an inverse relationship between flow rate and recovery. Hypertonic saline administered centrally caused an increase in dialysate and plasma oxytocin while the intravenous injection affected only plasma oxytocin. The excitatory amino acid, glutamate (0.05-0.5 M), caused an increase in plasma, but not dialysate oxytocin, while depolarization with potassium chloride (0.05-0.15 M) had no significant effects. Histological examination showed that the dialysis probe was located in the rostral, lateral PVN. Our results show that in vivo microdialysis provides a method for the delivery of drugs into specific brain regions as well as a useful technique for the evaluation of in vivo neuropeptide release.     

Estrogen-induced efflux of endogenous catecholamines from the hypothalamus in vitro

Short-term organ cultures of the intact hypothalamus were used to study the effects of various estrogenic compounds on catecholamine release. Estradiol-17 beta (0.1--20 microM) produced a concentration-dependent efflux of norepinephrine and dopamine while its biologically inactive enantiomer, estradiol-17 alpha, was ineffective at concentrations up to 20 microM. Diethylstilbestrol, a potent non-steroidal estrogen, was as effective as estradiol-17 beta in inducing catecholamine efflux. In contrast, weakly or non-estrogenic steroids such as estrone, estriol, and corticosterone were without effect. The time course of the estrogen-induced efflux of hypothalamic catecholamines was similar to that previously reported for the estrogen-induced accumulation of hypothalamic cAMP, providing further evidence for the involvement of catecholamines in this effect. Theses results suggest that estrogen may facilitate the release of catecholamines within the hypothalamus.     

The effects of four days of continuous striatal microdialysis on indices of dopamine and serotonin neurotransmission in rats

The effects of 4 days of continuous microdialysis with a small-diameter concentric-style probe on indices of striatal dopamine (DA) and serotonin neurotransmission were assessed. It was found that over 4 days of dialysis, there was a marked time-dependent decrease in the basal concentrations of 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in dialysate and in amphetamine-stimulated DA release. In contrast, there was no decrease in basal DA or in the ability of cocaine to elevate the concentration of DA in dialysate over the same period of time. There were only very modest changes in dialysate levels of the serotonin metabolite, 5-hydroxyindoleacetic acid (5-HIAA), relative to the marked changes in DA metabolites. It is suggested that 4 days of continuous dialysis does not result in a non-specific decrease in diffusibility of these compounds into the dialysis probe, but that the changes are more likely due to probe-induced damage to the nigrostriatal DA system. It is also suggested that a “stable” basal concentration of DA in dialysate is an especially poor indicator of the integrity of the dopaminergic input to the striatum. The implications of these findings for within-subjects design microdialysis experiments are discussed.     

Effects of two volatile anesthetics (sevoflurane and halothane) on the hypothalamic noradrenaline release in rat brain

There are marked increases in noradrenaline (NA) release during emergence from general anesthesia induced with volatile anesthetics. These changes in NA in the posterior hypothalamus of the rat were assessed by intracranial microdialysis. Sevoflurane and halothane in equipotent concentrations were used to obtain the same depth of anesthesia. NA release increased similarly with the two agents during recovery. However, NA release remained elevated longer with halothane, from which recovery was also slower.     

Vasopressin release within the supraoptic and paraventricular nuclei of the rat brain: osmotic stimulation via microdialysis.

The combination of microdialysis and a highly sensitive radioimmunoassay was used in order to monitor the in vivo release of arginine vasopressin (AVP) within hypothalamic supraoptic (SON) and paraventricular (PVN) nuclei of the rat brain. A dialysis probe was inserted into the SON or PVN area and microdialysis was performed in conscious or urethane-anesthetized animals before, during and after hypertonic artificial cerebrospinal fluid (aCSF, with 1 M NaCl) was delivered via the probe. The recovery of AVP in vitro was 1.60%, that of [3H]OH in vitro 14.2% and in vivo 8.44% (SON) and 9.26% (PVN), respectively. AVP was consistently detected in both SON and PVN dialysates; basal levels averaged 0.87 +/- 0.22 pg/30-min dialysate (SON, n = 51) and 0.80 +/- 0.24 pg/30-min dialysate (PVN, n = 6), respectively. Hypertonic aCSF given over a period of 30 min, 60 min or 90 min, resulted in an increased AVP release within the SON which, however, reached its peak (to 8.86-10.27 pg/sample; P less than 0.001 as compared to basal) only in the poststimulation period, i.e. after replacement of hypertonic with isotonic aCSF. An identical osmotic stimulus given 150-210 min after the first one produced similar, though slightly declined, changes in AVP release. In the PVN, AVP release patterns prior to and in response to the first hypertonic pulse were similar to those in the SON; a possible functional difference between the two nuclei is indicated by the lack of a rebound increase in AVP release following the second stimulation. The physiological significance of intranuclearly released AVP remains to be shown.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neurochemical evidence for inflammation-induced activation of the coeruleospinal modulation system in the rat

By using the microdialysis technique, the concentration of noradrenaline (NA) in the dorsal horn during unilateral hindpaw inflammation was compared between rats receiving bilateral lesions of the locus coeruleus (LC) and non-operated control rats. Bilateral lesions of the LC were made using an anodal current one week before testing. Unilateral hindpaw inflammation was produced by a subcutaneous injection of carrageenan (6 mg in 0.15 ml saline). Under conditions of sodium pentobarbital anesthesia, the microdialysis probe was inserted into the dorsal horn either ipsilateral or contralateral to the site of inflammation. The NA concentration in the dialysate was measured by high-performance liquid chromatography with electrochemical detection. Prior to carrageenan injection, the NA level (baseline level) did not differ between the LC-lesioned and the non-operated groups. After carrageenan injection, in the non-operated rats, the NA level increased significantly compared to the baseline level only in the dorsal horn ipsilateral to the site of inflammation, but not in the dorsal horn contralateral to the site of inflammation. An increase of the NA level was not observed in the LC-lesioned rats and in rats receiving an injection of saline. The result suggests that unilateral hindpaw inflammation produces excitation of descending NA-containing neurons from the LC, resulting in an increase of the NA level in the dorsal horn ipsilateral to the site of inflammation.     

Noradrenaline stimulates somatostatin release from incubated slices of the amygdala and the hypothalamic preoptic area

Neurotransmitter effects were studied on in vitro release of immunoreactive somatostatin (SRIF) from slices prepared from several regions of the rat brain: mediobasal hypothalamus (MBH), preoptic anterior hypothalamic area (POA) and amygdaloid complex (AMY). Potassium (K+, 56 mM) stimulated SRIF release in all structures tested in a calcium dependent manner. Morphine, dopamine, GABA and serotonin did not modify SRIF release in any structure; noradrenaline (NA) was not effective on MBH slices, but elicited a dose-dependent stimulation of SRIF release from POA and AMY (ED50 = 6.4 +/- 1.4 nM and 3.6 +/- 1.2 nM respectively). Converse orders of potency of adrenergic agonists were observed in both structures (POA, adrenaline greater than noradrenaline greater than isoproterenol; AMY, isoproterenol greater than adrenaline greater than noradrenaline). Phentolamine blocked NA-induced SRIF release in the POA while propranolol was ineffective. On the contrary, propranolol, but not phentolamine, antagonized NA stimulation in the amygdala. The data suggest that NA acting through specific receptors modulate SRIF release from POA and AMY. In POA, NA effect seems mediated through alpha adrenergic receptors while in AMY, beta receptors are involved. The possibility that these interactions of NA with SRIF release are correlated with effects of NA on growth hormone secretion or on epileptic events is discussed.     

Evidence for a cholinergic influence on catecholaminergic pathways terminating in the anterior and medial basal hypothalamus

Adult male Sprague-Dawley rats were given hourly injections of physostigmine for 1–4 h, and the effect of this treatment on dopamine (DA) and noradrenaline (NA) content or on DA and NA turnover was determined in the anterior hypothalamus, medial basal hypothalamus and telencephalon-thalamus. The turnover of DA and NA was estimated by measuring the decline in these amines produced following the inhibition of tyrosine hydroxylase with α-methyl-p-tyrosine (αMPT). In later experiments oxotremorine was administered instead of physostigmine at hourly intervals for 2 h. Physostigmine administration resulted in a highly significant increase in the depletion of NA produced by αMPT indicating that the turnover of NA was increased by this drug. This effect was observed in the medial basal hypothalamus and anterior hypothalamus but not in the telencephalon-thalamus. Oxotremorine also produced an increase in NA turnover, but this drug was effective in all three brain areas. Atropine pretreatment blocked the effect of both physostigmine and oxotremorine on NA turnover. However, in the case of physostigmine, atropine was only effective if it was given 30 min before each injection of physostigmine. Mecamylamine, a nicotine blocker, did not reverse the effect of physostigmine on NA turnover. These results suggest that there is a cholinergic input via muscarinic receptors which influences the activity of noradrenergic pathways terminating in the anterior or medial basal hypothalamus.     

Extracellular oxytocin in the paraventricular nucleus: hyperosmotic stimulation by in vivo microdialysis

The effect of central osmotic stimulation on oxytocin (OT) secretion from the paraventricular nucleus (PVN) was examined using a newly developed in vivo microdialysis technique. A dialysis probe was inserted into the PVN region and microdialysis was performed in conscious animals. Hyperosmotic solutions were delivered via the dialysis probe, and perfusate and blood samples were collected. OT was consistently detected in the PVN dialysate. Hyperosmotic sodium chloride (1 M) produced a significant increase in dialysate and plasma OT, whereas D-mannitol (2 M) had no effect. These results suggest that (1) in vivo microdialysis may provide a useful technique for the evaluation of neuropeptide secretion from specific brain regions and (2) there are sodium-sensitive cells in the PVN region which respond to increases in extracellular sodium, resulting in an increase in central and peripheral oxytocin secretion.     

Presynaptic regulation of the release of catecholamines in the cat hypothalamus

The posterior hypothalamus of cats was superfused through a push-pull cannula and the release of endogenous catecholamines was determined in the superfusate. Superfusion with yohimbine, isoprenaline, salbutamol or tazolol increased, while superfusion with propanolol decreased, the release of all three catecholamines. Transection of the brain caudal to the hypothalamus inhibited ‘resting’ and drug-induced release. It is concluded that α- and β-adrenoceptors of the hypothalamus are involved in the regulation of the release of catecholamines.     

Either desipramine or TMB-8 suppresses cyanide-induced norepinephrine efflux from in vivo cardiac sympathetic nerves of cats

To investigate the effect of hypoxia on endogenous norepinephrine (NE) release from cardiac sympathetic nerve ending, we administered sodium cyanide (NaCN) for 30 min into the myocardial interstitial space through a dialysis probe and measured dialysate NE levels. During the NaCN perfusion, a marked and concentration-dependent increase in dialysate NE was observed. This cyanide-induced NE response was suppressed by pretreatment with despiramine (a membraneous NE transport inhibitor). Furthermore, the cyanide-induced NE response was suppressed by pretreatment with TMB-8 (intracellular Ca(2+) antagonist) but unaffected by omega-conotoxin GVIA (NE releasing inhibitor). Our data suggest that two (desipramine or TMB-8 suppressive) mechanisms contributed to the amount of NE efflux induced by cyanide in in vivo cardiac sympathetic nerve.     

Central carbachol stimulates vasopressin release into interstitial fluid adjacent to the paraventricular nucleus

We have used an in vivo double microdialysis probe technique in conscious rats to determine whether the application of carbachol to one paraventricular nucleus (PVN) can result in increased local release of vasopressin from that PVN. Experiments were carried out 24 h after placement of microdialysis probes lateral to each PVN. When both probes were perfused initially with 0.9% NaCl, vasopressin was detected in the outflow (dialysate) from both probes. When carbachol (100 μg/ml) was included in the perfusate of one probe for the first 10 min of a 30-min collection period, while the other probe continued to be perfused with saline alone, there was a seven-fold increase in the concentration of vasopressin in the dialysate from the carbachol-perfused probe; the vasopressin concentration in the dialysate from the contralateral probe increased only slightly. The plasma vasopressin concentration was also elevated. When one of the paired probes was perfused with carbachol (100 μg/ml) for 30 min, there were similar increases in the concentration of vasopressin in the dialysate from both probes and a sustained increase in the plasma vasopressin concentration. Thus, vasopressin is released into the interstitial fluid adjacent to the PVN under basal conditions, and this release can be substantially increased when vasopressin secretion to the periphery is stimulated.     

Local epinephrine release in the rabbit myocardial interstitium in vivo

Although several investigations have suggested cardiac epinephrine (Epi) release, local Epi release in the myocardial interstitium in vivo has not been measured. Using cardiac microdialysis in the rabbit, we measured dialysate Epi and norepinephrine (NE) concentrations as indices of myocardial interstitial Epi and NE levels, respectively. Exocytotic release induced by local administration of KCl (100 mM) through the dialysis probe increased Epi to 24.2 +/- 13.2 pg/ml from a control value of 3.2 +/- 3.6 pg/ml (P < 0.01, n = 6). Non-exocytotic release induced by the local administration of tyramine (10 microg/ml) also increased Epi to 34.6 +/- 15.3 pg/ml (p < 0.05 from control, n = 6). We conclude that Epi can be released via both exocytotic and non-exocytotic release mechanisms from the heart.     

Variations in extracellular monoamines in the prefrontal cortex and medial hypothalamus after modafinil administration: a microdialysis study in rats.

The role of brain amines in mediating the effects of the wake-promoting agent modafinil, used in the treatment of sleepiness associated with narcolepsy is still uncertain. Therefore we studied the effects of modafinil on extracellular serotonin (5-HT), dopamine (DA) and noradrenaline (NA), in rat prefrontal cortex and in the medial hypothalamus area. Modafinil (128 mg/kg i.p.) significantly increased waking in the first 4 h of EEG sleep recording. This cortical and behavioral activation was associated with an initial increase in extracellular 5-HT, DA and NA during the first 60 min following modafinil administration. In the prefrontal cortex, 5-HT release remained high for 3 h after modafinil administration. In contrast, in the hypothalamus, only NA release was enhanced while DA and 5-HT levels remained low. In a first step, modafinil may generate waking partly via cortical monoamine release, particularly DA and 5-HT, and also hypothalamic NA. In a second step, maintenance of waking might depend on hypothalamic NA.