Microinjection of carbachol in the lateral hypothalamus produces opposing actions on nociception mediated by α1- and α2-adrenoceptors

Electrical stimulation of the lateral hypothalamus (LH) produces antinociception partially blocked by intrathecal α-adrenergic antagonists, but the mechanism underlying this effect is not clear. Evidence from immunological studies demonstrates that substance P-immunoreactive neurons in the LH project near the A7 catecholamine cell group, a group of noradrenergic neurons in the pons known to effect antinociception in the spinal cord dorsal horn. Such evidence suggests that LH neurons may activate A7 neurons to produce antinociception. To test this hypothesis, the cholinergic agonist carbachol was microinjected into the LH at doses of 63, 125 and 250 nmol and the resulting effects on tail-flick and nociceptive foot-withdrawal latencies were measured. All three doses significantly increased response latencies on both tests, with the 125-nmol dose providing the optimal effect. Intrathecal injection of the opioid antagonist naltrexone (97 nmol) partially reversed antinociception, but neither the α₂-adrenoceptor antagonist yohimbine nor the α₁-adrenoceptor antagonist WB4101 altered latencies. However, two sequential doses of yohimbine blocked LH-induced antinociception on both tests. In contrast, two sequential doses of WB4101 increased nociceptive responses on both the tail-flick and foot-withdrawal tests. These findings, and those of published reports, suggest that neurons in the LH activate spinally projecting methionine enkephalin neurons, as well as two populations of A7 noradrenergic neurons that exert a bidirectional effect on nociception. One of these populations increases nociception through the action of α₁-adrenoceptors and the other inhibits nociception through the action of α₂-adrenoceptors in the spinal cord dorsal horn.     

Central α1 adrenoceptor subtypes in narcolepsy-cataplexy: a disorder of REM sleep

The present study suggests the specific involvement within the central nervous system of an α₁ adrenoceptor subtype in a behavior, the control of cataplexy, a pathological analogue of rapid eye movement (REM) sleep atonia. Experiments have shown that prazosin, an α₁ antagonist, dramatically aggravates canine narcolepsy-cataplexy through a central mechanism, and that [ ³H]prazosin binding sites are increased in the amygdala of narcoleptic dogs¹⁶. However, the corresponding Scatchard plots were curvilinear and best fit was obtained with a two-site model, suggesting the existence of two [ ³H]prazosin binding sites¹⁸. These two sites (high and low affinity [ ³H]prazosin binding sites) met the criteria for authentic receptors and were respectively very similar to the α_1a an α_1b (high and low affinity for WB4101, respectively) subtypes recently described in the rat and rabbit. Our results of in vivo pharmacology and in vitro [ ³H]prazosin binding in canine narcolepsy now clearly implicate the low affinity [ ³H]prazosin binding site (α_b) in canine narcolepsy: (1) Prazosin, an α₁ antagonist with similar affinity for both subtypes, was much more potent in increasing cataplexy than WB4101, a compound with more affinity for the α_1a receptor. (2) Chlorethylclonidine and phenoxybenzamine, two irreversible blockers of the α₁ receptors with more affinity for the α_1b receptors, aggravate cataplexy for up to two weeks. (3) The α₁ receptor upregulation previously reported by our group in the amygdala of narcoleptic dogs was due to a selective increase in the low affinity [ ³H]prazosin binding sites. A role for noradrenaline in REM sleep regulation has been suspected for many years, but has never been clearly elucidated. Our data provide evidence for its specific involvement through a specific central α₁-adrenergic receptor subtype.     

Relationships between β- and α2-adrenoceptors and G coupling proteins in the human brain: effects of age and suicide

Interactions between brain α₂- and β-adrenoceptors are of interest in physiological (aging) and pathological (major depression) processes involving both receptors. In this study, total β-adrenoceptors and β_1/2-subtypes were quantitated in postmortem human brains to investigate their relationships with α_2A-adrenoceptors and specific G proteins during the process of aging and in brains of suicide victims. Analysis of [³H]CGP12177 binding, in the presence of CGP20712A (β₁-antagonist), indicated that the predominant β-adrenoceptor in the frontal cortex is the β₁-subtype (65–75%). The density of total β- (r=−0.60, n=44) or β₁-adrenoceptors (r=−0.78, n=22), but not the β₂-subtype, declined with aging (3–80 years). The density of total β- or β₁-adrenoceptors, but not the β₂-subtype, correlated with the number of α₂-adrenoceptors quantitated in the same brains with the agonist [³H]UK14304 (r=0.71–0.81) or the antagonist [³H]RX821002 (r=0.61–0.66). Interestingly, the ratios α₂/β- or α₂/β₁-adrenoceptors did not correlate with the age of the subject at death, indicating that the proportion of α₂/β-adrenoceptors in brain remains rather constant during the process of aging. The density of β-adrenoceptors correlated with the immunodensity of Gαs (r=0.55) and Gβ (r=0.61) proteins, and that of α₂-adrenoceptors with those of Gαi_1/2 (r=0.88) and Gβ (r=0.65). In brains of suicides, compared to controls, the ratio between α₂- and β- or β₁-adrenoceptors (α₂-full agonist sites/β-sites) was greater (1.3- to 2.0-fold; P<0.05). The results demonstrate a close interdependence between brain α₂- and β-adrenoceptors during aging, and in brains of suicides. The quantitation of the α_2A/β-adrenoceptor ratio could represent a relevant neurochemical index in the study of brain pathologies in which both receptors are involved.     

Regulation of α2A-adrenoceptor expression by chronic stress in neurons of the brain stem

α₂-Adrenoceptors are supposed to be important regulatory elements in responses to stress. Previous receptor binding studies in male tree shrews have shown that chronic psychosocial stress down-regulates binding sites for α₂-adrenergic ligands in several brain stem nuclei. The aim of the present study was to quantify effects of chronic subordination stress on expression of the α₂-adrenoceptor subtype A gene in identified neurons of the brain stem. We partially cloned the α_2A-adrenoceptor cDNA of the tree shrew (1.22 kb) and localized receptor RNA expression in brain stem neurons by in situ hybridization using a ³⁵S-labeled cRNA probe (1.06 kb). To identify neurons expressing receptor mRNA, brain sections were first immunocytochemically stained with antibodies against tyrosine hydroxylase, phenylethanolamine-N-methyltransferase, or glutamate, and then processed for in situ hybridization. Furthermore, expression of receptor-specific RNA was quantified in single neurons of animals which had been psychosocially stressed during 4 weeks and in unstressed controls. We found strong in situ hybridization in the noradrenergic neurons of the locus coeruleus, but only weak labeling of A2 neurons in the solitary tract nucleus and no labeling of A1 neurons in the caudal ventrolateral medulla. Adrenergic neurons in the solitary tract nucleus (group C2) did not express the α_2A-adrenoceptor, and C1 neurons in the rostral ventrolateral medulla showed only a minor labeling by the in situ probe. In contrast, large glutamatergic neurons in the lateral reticular nucleus were strongly labeled by the probe. Chronic psychosocial stress reduced α_2A-adrenoceptor RNA expression in locus coeruleus neurons (−24.0%), in solitary tract neurons (−31.0%), and in neurons of the lateral reticular nucleus (−18.8%). These findings show that stress not only decreases the expression of the α_2A-adrenergic autoreceptor in the locus coeruleus but also of α_2A-heteroreceptors in glutamatergic neurons.     

Spatial working memory improvement by an α2-adrenoceptor agonist dexmedetomidine is not mediated through α2C-adrenoceptor

1. Aged α_2C-adrenoceptor knockout and wild type mice were used to investigate whether α_2C-adrenoceptors are involved in mediating the beneficial effects of α₂-adrenoceptor agonist, dexmedetomidine, on spatial working memory.

2. A win-stay task in the radial arm maze was used to dissociate the effects of dexmedetomidine on working vs. reference memory. In addition, the animals were tested in simple response habit learning in the T-maze.

3. Knockout mice made more working memory errors after the change of the baited arm in radial arm maze, but after training reached again as accurate level of performance as wild type controls. Dexmedetomidine 5 and 10 μg/kg alleviated the increase in spatial working memory errors after the change of the baited arm in knockout mice. Knockout and wild type mice performed equally well in T-maze, and dexmedetomidine had no effect on this simple response learning.

4. The present results indicate that α₂-adrenoceptor agonists have a selective effect on spatial working memory not only in monkeys but also in mice. Further, this study confirms our earlier finding that the presence of α_2C-adrenoceptors is not necessary for the spatial working memory enhancing effect of α₂-adrenoceptor agonists.

The resolution of dopamine and β1- and β2-adrenergic-sensitive adenylate cyclase activities in homogenates of cat cerebellum, hippocampus and cerebral cortex

The stimulation of adenylate cyclase by dopamine and various β-adrenergic agonists has been investigated in homogenates from 3 areas of cat brain: the cerebral cortex, cerebellum and hippocampus. The purpose of the study was to determine whether the β-adrenergic receptors coupled to adenylate cyclase could be classified as either β₁ and β₂ subtypes in the different regions studied.The stimulation of adenylate cyclase by the β-adrenergic agonist, (−)isoproterenol (5 × 10⁻⁶M), was completely blocked by the specific β-adrenergic antagonist, (−)alprenolol (10⁻⁵ M), but not by the dopaminergic antagonist, fluphenazine (10⁻⁵ M), whereas the stimulation of adenylate cyclase by (−)epinephrine (10⁻⁴ M) was blocked to varying extents by these two drugs in each of the 3 regions studied. The (−)epinephrine effect was always blocked in the combined presence of (−)alprenolol and fluphenazine. The adenylate cyclase stimulation by (−)epinephrine which is not blocked by (−)alprenolol was due to interaction of (−)epinephrine with a dopaminergic-sensitive adenylate cyclase which has been characterized in cerebral cortex, hippocampus and cerebellum.Regional differences in the affinity of β-adrenergic-sensitive adenylate cyclase for various agonists were investigated in the presence of fluphenazine (10⁻⁵ M). In the cerebellum the potency order was (±)protokylol> (±)hydroxybenzylisoproterenol> (±)isoproterenol> (−)epinephrine> (±)salbutamol> (−)norepinephrine, indicating the presence of a β₂-adrenergic receptor. In the cerebral cortex the potency order was (−)isoproterenol> (±)protokylol> (±)hydroxybenzylisoproterenol> (−)epinephrine= (−)norepinephrine((±)salbutamol being inactive). A similar pattern was found in the hippocampus indicating the presence of a β₁-adrenergic receptor in these two regions. (±)Salbutamol was a partial agonist in the cerebellum and a competitive antagonist in the cerebral cortex.The ratio of the antagonist potencies of (±)practolol and (±)butoxamine preferential β₁- and β₂-adrenergic antagonists respectively, to block the stimulation of adenylate cyclase was 25 in the cerebellum, compared to 0.5 in the cerebral cortex and 1.6 in the hippocampus. These results confirm the presence of a β₂ subtype of receptor coupled to adenylate cyclase in the former and β₁ subtypes in the latter two regions. The comparison between the affinities of a series of β-adrenergic agonists and antagonists for the β-adrenergic receptors coupled with an adenylate cyclase in cerebral cortex and cerebellum with their affinities for well characterized β₂-adrenergic receptors in lung and β₁-adrenergic receptor in heart substantiated this conclusion.     

Evidence for central α2-adrenergic modulation of rat pineal melatonin synthesis

This study was performed to distinguish central and peripheral α₂-adrenoceptors in the inhibition of rat pineal melatonin synthesis. The rats received lipo- or hydrophilic α₂-adrenoceptor ligand injections at middark; after 1 or 2 h the pineal melatonin contents were measured. The lipophilic agonist medetomidine (100 μg/kg s.c.) suppressed the melatonin contents significantly, while the hydrophilic agonists ST-91 and p-aminoclonidine (10 or 100 μg/kg i.v.) did not. The suppression by medetomidine was counteracted by the lipophilic antagonist yohimbine (0.3–3.0 mg/kg i.p.) but not by the hydrophilic antagonist L-659,066 (1–10 mg/kg i.v.). In conclusion, the suppression of nocturnal melatonin synthesis by α₂-adrenoceptor agonists is mainly of central origin.     

Noradrenaline- and time-dependent changes in neocortical α2- and β1-adrenoceptor binding in dorsal noradrenergic bundle-lesioned rats

A 6-hydroxydopamine-induced lesion of the dorsal noradrenergic bundle (DNB) in rats markedly decreased neocortical noradrenaline concentration (NNC) by 72–100% as measured 1, 3 and 13 months after the lesioning procedure. The concomitant assessment of neocortical α₂- and β₁-adrenoceptor binding (NAAB and NBAB, respectively) usually indicated significant increases of 25–74% for these two variables. There were, however, cases of unchanged NAAB and NBAB which presumably reflected an incomplete DNB lesion and a consequent time-related, partial recovery of NNC. The results emphasize the potential for long-term sequelae of the DNB lesion, and the existence of a critical NNC threshold (10–30% of control NNC values) which modulates postsynaptic α₂ and β₁-adrenoceptor density.     

Autoradiographic studies of central α2A- and α2C-adrenoceptors in the rat using [H]MK912 and subtype-selective drugs

In the present study we examined the distribution of α_2A- and α_2C-adrenoceptors in tissue slices from the rat cervical spinal cord and from brain slices collected at the level of the striatum. To differentiate between α_2A- and α_2C-adrenoceptors, the slices were incubated with [³H]MK912 in the presence of graded concentrations of the α_2A-selective drug, BRL44408, or the α_2C-selective drug, spiroxatrine. Computer analysis of the autoradiograms indicated that 0.4 nM [³H]MK912 plus 185 nM BRL44408 selectively labeled α_2C-adrenoceptors, while 0.4 nM [³H]MK912 plus 220 nM spiroxatrine selectively labeled α_2A-adrenoceptors. Using this approach, α_2C-adrenoceptors were detected in the striatum, while α_2A-adrenoceptors predominated in the cortical layers 1–4, the spinal cord distal dorsal horn, the septum and the endopiriform nucleus.     

Subtype-specificity of the presynaptic α2-adrenoceptors modulating hippocampal norepinephrine release in rat

In vivo brain microdialysis and high-performance liquid chromatography with electrochemical detection were used to study the effect of different selective α₂-antagonists on hippocampal norepinephrine (NE) release in freely moving awake rat. Systemic administration (0.5 mg/kg i.p.) of either the α_2AD-antagonist BRL 44408 or the α-_2BC-antagonist ARC 239 did not significantly change the basal release of NE. At a higher dose (5 mg/kg i.p.) ARC 239 was still ineffective, whereas BRL 44408 caused a significant increase of the extracellular level of NE. Similar results were obtained from in vitro perfusion experiments. Rat hippocampal slices were loaded with [³H]NE and the electrical stimulation-evoked release of [³H]NE was determined. The α₂-antagonists were applied in a concentration range of 10⁻⁸ to 10⁻⁶ M. ARC 239 was ineffective, whereas BRL 44408 significantly increased the electrically induced release of [³H]NE. In agreement with the data of microdialysis and perfusion experiments. BRL 44408 displaced [³H]yohimbine from hippocampal and cortical membranes of rat brain with high affinity whereas ARC 239 was less effective. The pK_i values of eight different α₂-adrenergic compounds showed a very good correlation (r = 0.98, slope = 1.11 P < 0.0001) in hippocampus and frontal cortex where the α₂-adrenoceptors have been characterized as α_2D-subtype. Our data indicate that hippocampal NE release in rat is regulated by α_2D-adrenoceptors, a species variation of the human α_2A-subtype.     

Inhibition of interleukin-1β and prostaglandin E2 thermogenesis by glycyl-glutamine, a pro-opiomelanocortin-derived peptide

Interleukin-1β (IL-1β) and other cytokines produce fever by stimulating prostaglandin E₂ (PGE₂) synthesis in thermoregulatory regions of the preoptic area and anterior hypothalamus (POA/AH). Prostaglandin E₂ is thought to raise body temperature, at least in part, by stimulating β-endorphin release from pro-opiomelanocortin neurons that innervate the POA/AH. In this study, we investigated whether glycyl-glutamine (β-endorphin_30–31), an inhibitory dipeptide synthesized from β-endorphin post-translationally, inhibits IL-1β and PGE₂-induced hyperthermia. Hyperthermic sites were identified by microinjecting PGE₂ (3 fmol/1 μl) into the medial preoptic area (mPOA) of conscious, unrestrained rats. Interleukin-1β (1 U) injection into the same PGE₂ responsive thermogenic sites in the mPOA elicited a prolonged rise in colonic temperature (T_c) (+1.02±0.06°C) that persisted for at least 2 h. Glycyl-glutamine (3 nmol) co-injection into the mPOA inhibited IL-1β thermogenesis completely (T_c=−0.18±0.22°C). Glycyl-glutamine had no effect on body temperature when given alone to normothermic rats. Co-injection of individual amino acids, glycine and glutamine (3 nmol each amino acid), failed to influence IL-1β-induced thermogenesis, which indicates that Gly-Gln hydrolysis does not explain its inhibitory activity. Glycyl-glutamine (3 nmol) also prevented the rise in body temperature produced by PGE₂ (PGE₂=0.89±0.05°C; PGE₂ plus Gly-Gln=−0.16±0.14°C), consistent with evidence that PGE₂ mediates IL-1β-induced fever. These findings demonstrate that Gly-Gln inhibits the thermogenic response to endogenous pyrogens.     

Characteristics of the β1- and β2-adrenergic-sensitive adenylate cyclases in glial cell primary cultures and their comparison with β2-adrenergic-sensitive adenylate cyclase of meningeal cells

The agonist specificity pattern of the β-adrenergic adenylate cyclase in glial primary cultures was not typical of either β₁- or β₂-adrenergic receptors. The dose-response curves for adrenaline did not correspond to simple mass action kinetics and their computer analysis suggests the presence of both β₁- and β₂-adrenergic-sensitive adenylate cyclase (58 ± 17% and 42 ± 17% respectively).Similar properties of β₁- and β₂-adrenergic-sensitive adenylate cyclases were found by computer analysis of the dose-response curves for isoprenaline in the presence of a constant concentration of practolol (a selective β₁ antagonist) ( 55 ± 10% and 45 ± 10% of β₁- and β₂-sensitive adenylate cyclase respectively).The curves for displacement of [³H]dihydroalprenolol by practolol confirm these results.For purpose of comparison, the β-adrenergic receptors of meningeal cells in cultures were subjected to similar analysis. The results clearly showed that these cells exclusively contained β₂-adrenergic receptors.     

Inhibition of spontaneous inhibitory postsynaptic currents (IPSC) by noradrenaline in rat supraoptic neurons through presynaptic α2-adrenoceptors

It has been shown that noradrenergic activation has great influence on the activities of hypothalamic supraoptic neurons. No direct evidence has been reported on the presynaptic effects of adrenoceptors in the actions of noradrenaline on supraoptic neurons, although postsynaptic mechanisms have been studied extensively. In the present study, we explored presynaptic effects of noradrenaline on the supraoptic neurons by measuring spontaneous inhibitory postsynaptic currents (IPSC) with the whole-cell patch-clamp technique. Noradrenaline reduced the frequency of IPSCs in a dose-dependent (10⁻⁹ to 10⁻³ M) and reversible manner. Noradrenaline did not affect the amplitude of IPSCs at concentrations of 10⁻⁹ to 10⁻⁵ M, but reduced the amplitude of IPSCs at high concentrations (10⁻⁴ and 10⁻³ M). The inhibitory effects of noradrenaline were mimicked by the α₂-agonist clonidine (10⁻⁴ M), but not by the α₁-agonist methoxamine (10⁻⁴ M) nor by the β-agonist isoproterenol (10⁻⁴ M). Moreover, the inhibitory effects of noradrenaline on IPSCs were blocked by the non-selective α antagonist phentolamine (10⁻⁴ M) or the selective α₂-antagonist yohimbine (10⁻⁴ M), but not by the α₁-antagonist prazosin (10⁻⁴ M). These results suggest that noradrenaline inhibits release of GABA from the presynaptic GABAergic terminals of the supraoptic neurons by activating presynaptic α₂-adrenoceptors and such presynaptic mechanisms may play a role in the excitatory control of SON neurons by noradrenergic neurons.     

Increased β2-adrenoceptors in the superior cervical ganglia of genetically hypertensive rats

[¹²⁵I]Iodocyanopindolol binding sites were characterized by autoradiography in the superior cervical ganglia of Wistar-Kyoto (WKY) rats. A high concentration of (−)-[¹²⁵I]iodocyanopindolol binding sites, characterized as β-adrenoceptors by (−)-propranolol displacement, was distributed throughout the ganglia and in the postganglionic (internal carotid) nerve. ICI 118,551, a β₂-selective antagonist, displaced more than 85% of the binding sites, whereas CGP 20712A, a β₁-selective antagonist, displaced less than 10% of the binding sites, indicating that the β-adrenoceptors were primarily of the β₂-subtype. Emulsion autoradiography demonstrated that at least part of the binding sites were associated with principal ganglion cells. Unilateral deafferentation did not modify the number of binding sites in the superior cervical ganglia of WKY or spontaneously hypertensive rat (SHR). These results suggest that at least part of these receptors may correspond to prejunctional β₂-adrenoceptors originated in principal ganglion cells. The concentration of β₂-receptors was increased in the superior cervical ganglia of young and adult SHR when compared to age-matched WKY rats (49% and 39%, respectively). There were no differences in β₂-adrenoceptor number in the stellate ganglia of young and adult WKY and SHR. These results suggest that β₂-adrenoceptor stimulation may be selectively enhanced in some peripheral sympathetic ganglia in SHR and this could play a role in the development and maintenance of the increased symphatetic activity in this strain.     

Substance P suppresses the activity of α2-adrenoceptors of the nucleus reticularis gigantocellularis involved in cardiovascular regulation in the rat

We evaluated possible interactions between substance P (SP) and the α₂-adrenoceptors in the nucleus reticularis gigantocellularis (NRGC) of the medulla oblongata involved in cardiovascular regulation. Adult, male Sprague-Dawley rats anesthetized with pentobarbital sodium (40 mg/kg, i.p., with 10 mg/kg/h i.v. supplements) were used. The circulatory suppressant efficacy of a centrally acting α₂-adrenoceptor agonist, guanabenz, was used as the experimental index. Bilateral microinjection of SP (300 or 600 pmol) into the NRGC, a medullary site that is critically involved in the cardiovascular depressive actions of guanabenz, significantly diminished the hypotensive and bradycardiac efficacy of the aminoguanidine compound (100 μg/kg, i.v.). This implied reduction in α₂-adrenoceptor activity in the NRGC by SP was antagonized by its selective receptor antagonist, [d-Pro²,d-Trp^7,9]-SP (1200 pmol). Similarly, attenuation by SP of the cardiovascular suppressant effects of guanabenz was also reversed by immunocytochemically verified depletion of dopamine-β-hydroxylase-immunoreactive nerve terminals in the NRGC, elicited by the selective noradrenergic neurotoxin, DSP4 (50 μg). These data suggest that SP may exert an inhibitory action on the α₂-adrenoceptors in the NRGC that are involved in central cardiovascular regulation, possibly via a presynaptic modulation on noradrenergic neurotransmission.     

Hypothalamic α2- and β-adrenoceptor rhythms are correlated with circadian feeding: evidence from hronic methamphetamine treatment and withdrawal

The circadian regulation of food intake in rats is correlated with a bimodal rhythm of β-adrenoceptor binding in the lateral hypothalamus and a unimodal rhythm of α₂-adrenoceptor binding in the medial hypothalamus. Chronic methampheamine treatment provides evidence for a functional correlation: β-adrenoceptor binding in the lateral hypothalamus is reduced at dusk, together with reduction of food intake; α₂-adrenoceptor binding in the medial hypothalamus is increased at dawn, together with persistent food intake. Longterm changes in these two adrenergic systems are also correlated with homeostasis of food intakke: 24-h mean β-adrenoceptor binding is reduced and α₂-adrenoceptor binding is increased upon methamphetamine withdrawal, when rebound feeding occurs. Corticosterone, although normally coupled to adrenergic mechan isms that regulate feeding, is phase delayed after chronic methamphetamine treatment.     

Distribution of α-adrenergic, β-adrenergic and dopaminergic receptors in discrete hypothalamic areas of rat

Catecholamine receptor binding sites were measured in discrete hypothalamic nuclei or regions as well as in certain extrahypothalamic areas of the adult male rat. For each assay, discrete areas were microdissected from frozen tissue sections and pooled from several animals. Specific high affinity binding sites were assessed at fixed ligand concentrations for [³H]p-aminoclonidine (PAC) and [³H](2-C 2′,6′-(CH₃O)₂ phenoxyethylamino)-methylbenzodioxan (WB-4101) for α-adrenergic receptor sites, for [³H]dihydroalprenolol (DHA) for β-adrenergic receptor sites, and for [³H]2-amino-6, 7-dihydroxy-1,2,3,4-tetrahydronaphtalene (ADTN) and [³H]spiroperidol in the presence of cinanserin for dopaminergic receptor sites.Regional variations in [³H]WB-4101 binding were relatively small in magnitude, with most hypothalamic and extrahypothalamic areas possessing between 60 and 90% of the binding in frontal cortex. [³H]PAC binding showed a wider range of binding density across brain areas than did [³H]WB-4101, but, in general, variations in [³H]PAC binding paralleled those in [³H]WB-4101 binding. In hypothalamus, binding was characterized as being predominantly to α₁-receptors in the of [³H]WB-4101 and to α₂-receptors in the case of [³H]PAC. The medial hypothalamic areas exhibited a somewhat higher density of these α-adrenergic sites than did the lateral hypothalamus (perifornical hypothalamus and medial forebrain bundle). Also, the ratio of [³H]PAC to [³H]WB-4101 binding differed in different hypothalamic areas, ranging from 1.5:1 to 4:1. The median eminence was exceptional in that it contained appreciable [³H]PAC but no significant [³H]WB-4101 binding sites at the radioligand concentrations tested. Binding of [³H]DHA to β-adrenergic receptors varied over approximately a 3-fold range in the different hypothalamic areas, with binding highest in the medial forebrain bundle and the medial preoptic area, and lowest in the periventricular, dorsomedial and posterior hypothalamic nuclei, the median eminence and the zona incerta. The ratio of β-adrenergic to α-adrenergic binding sites was generally lower in the medial than in the lateral hypothalamic areas and higher in the extrahypothalamic areas examined than in the hypothalamus. With regard to [³H]spiroperidol and [³H]ADTN binding to dopaminergic sites, the striatum, nucleus accumbens and olfactory tubercle showed a greater density of [³H]spiroperidol than of [³H]ADTN sites, in contrast to the hypothalamus where [³H]ADTN binding was more predominant. Within the hypothalamus, [³H]ADTN binding was relatively uniform, while [³H]spiroperidol binding was quite high in four hypothalamic areas (lateral perifornical area, medial forebrain bundle, paraventricular and dorsomedial nuclei), intermediate in the median eminence and arcuate nucleus, and low or not detectable in all other hypothalamic areas.     

Quantitative light microscopic autoradiographic localization of α2-adrenoceptors in the human brain

In the present work the anatomical distribution of α₂-adrenoceptors in the human central nervous system was studied in detail by quantitative autoradiography using the selective α₂ agonist [³H]bromoxidine ([³H]UK-14304) as a ligand. Only postmortem tissues from subjects free of neurological disorders were used in this study. Very high or high densities of α₂-adrenoceptors were found along layers I and III in non-visual neocortex, layers III and IVc of the visual cortex, CA₁ field — stratum lacunosum-moleculare — and dentate gyrus — stratum granularis — at the hippocampal formation, nucleus arcuatus at the hypothalamus, locus ceruleus, nucleus dorsalis of vagus and at the stratum granularis of the cerebellar cortex. Relevant densities of α₂-adrenoceptors were also observed along the remaining layers of neocortex, nuclei centralis, medialis and corticalis at the amygdala, anterior thalamic group and rotundocellularis nuclei, paraventricular and ventromedial hypothalamic nuclei, substantia innominata, superior colliculus — stratum zonale — and lateral periaqueductal area at the midbrain, nucleus tractus solitarii and dorsal horn — substantia gelatinosa — of the spinal cord. [³H]Bromoxidine specific binding was very low or negligible in the remaining brain areas. Although a general parallelism between the distribution of these receptors could be observed for the rat and human brain, dramatic species differences in the level of α₂-receptors were found in several brain areas, such as thalamus, amygdala or cerebellar cortex. In general, the distribution of α₂-adrenoceptors in the human brain found here was parallel to that described for the noradrenergic presynaptic terminals in the mammalian central nervous system, lending some weight to the proposed predominant presynaptic localization of these receptors. The relevance of the anatomical distribution of α₂-adrenoceptors in the human brain for a better knowledge of the neurochemistry of neuropsychiatric disorders is discussed.     

The role of α2-adrenoceptors of the medullary lateral reticular nucleus in spinal antinociception in rats

We attempted to find out the role of α₂-adrenoceptors of the medullary lateral reticular nucleus (LRN) in antinociception in rats. Spinal antinociception was evaluated using the tail-flick test, and supraspinal antinociception using the hotplate test. Antinociceptive effects were determined following local electric stimulation of the LRN, and following microinjections of medetomidine (an α₂-adrenoceptor agonist; 1–10 μg), atipamezole (an α₂-adrenoceptor antagonist; 20 μg) or lidocaine (4%) into the LRN. The experiments were performed using intact and spinalized Hannover-Wistar rats with a unilateral chronic guide cannula. Electric stimulation of the LRN as well as of the periaqueductal gray produced a significant spinal antinociceptive effect in intact rats. Medetomidine (1–10 μg), when microinjected into the LRN, produced no significant antinociceptive effect in the tail-flick test in intact rats. However, following spinalization, medetomidine in the LRN (10 μg) produced a significant atipamezole-reversible antinociceptive effect in the tail-flick test in the hot-plate test, medetomidine (10 μg) in the LRN produced a significant atipamezole-reversible increase of the paw-lick latency in intact rats. Microinjection of atipamezole (20 μg) or lidocaine alone into the LRN produced no significant effects in the tail-flick test. The results are in line with the previous evidence indicating brat the LRN and the adjacent ventrolateral medulla is involved in descending inhibition of spinal nocifensive responses. However, α₂-adrenoceptors in the LRN do not mediate spinal antinociception but, on the contrary, their activation counteracts antinociception at the spinal cord level. The spinal aninociceptive effect of supraspinally administered medetomidine in spinalized rats can be explained by a spread of the drug (e.g., via circulation) which then directly activates α₂-adrenoceptors at the spinal cord level.     

Influence of centrally administered α- and γ2-melanocyte-stimulating hormone on hypothalamic blood flow autoregulation in the rat

The effect of intracerebroventricularly (i.c.v.) administered α-melanocyte-stimulating hormone (MSH) and γ₂-MSH on hypothalamic blood flow autoregulation was studied in anesthetized rats at different levels of standardized arterial hypotension. Autoregulation was impaired upon i.c.v. administration of 5 γ g/kg γ₂-MSH while α-MSH caused no change.. Since this effect of γ₂-MSH wa identical to that produced by i.c.v. naloxone in the same model, γ₂-MSH may be a functional antagonist of central opioid mechanisms participating in the control of cerebral blood flow autoregulation.