Modulatory effects of catecholamines on neurons of the rat visual cortex: single-cell iontophoretic studies

The catecholamines noradrenaline and dopamine have been proposed as neuromodulators of cortical neuron excitability, and such a regulation could be mediated by specific adrenergic and dopaminergic receptors. We characterized electrophysiologically some of the types of responses to the iontophoretic application of adrenergic and dopaminergic agonists and antagonists on single cells in the rat visual cortex (areas occipital 1 monocular or Oc1M and occipital 1 binocular or Oc1B). For the majority of spontaneously active and visual cortical cells, noradrenaline and dopamine decreased the firing frequency. In the case of visually driven (synaptically activated) neurons, background firing was the main component of the response to be inhibited by the administration of noradrenaline, clonidine, and oxymetazoline, leading to an enhancement of the signal-to-noise ratio. Since these effects could be reduced or blocked by a previous ejection of the specific alpha 2-antagonist idazoxan, the findings support a role for alpha 2-adrenergic receptors in the transmission of sensory inputs to the visual cortex. These effects were not found with the mixed alpha-adrenergic agonist phenylephrine nor with the beta-agonist isoproterenol. Finally, the use of the inhibitory amino acid GABA rules out a simple hyperpolarizing response as the mechanism underlying noradrenaline modulatory effects in the cerebral cortex.     

The effect of pertussis toxin on alpha-2-adrenoceptor-mediated pigment migration in fish melanophores

The aggregation of melanin-granules within fish pigment cells (melanophores) can be elicited either by electrical stimulation of intrinsic nerves or by the addition of adrenergic agonists. The pigment aggregation seems to be mediated by alpha-2-adrenoceptors. In this investigation we have used various agonists and antagonists (noradrenaline, (+)- and (-)-adrenaline, isoprenaline, yohimbine and prazosin) to further characterize the pigment-aggregating receptor of Labrus ossifagus. All the results obtained support the notion of alpha-2-adrenoceptor-mediated pigment aggregation. The pertussis toxin, islet-activating protein (IAP), is known to inhibit the alpha-2-adrenoceptor-mediated signal transduction in mammals. We have used IAP to investigated whether fish melanophore alpha-2-adrenoceptors are also inhibited by this toxin. We found that IAP inactivated the alpha-2-adrenoceptor-mediated pigment aggregation in a dose-dependent manner. The inhibitory IAP-effect had a remarkably short onset-time in the melanophores (maximal effect was obtained within 10 min of incubation). Interestingly, binding of an agonist (noradrenaline) to the receptors prevented IAP from exerting its inhibitory action, whereas binding of an antagonist (yohimbine) gave no protection against the IAP-inactivation. In conclusion, the pigment-aggregating receptors of melanophores of L. ossifagus are very similar to the mammalian alpha-2-adrenoceptors. It is possible to inactivate the melanophore receptor system with IAP and this inactivation has a remarkably short onset-time. Stimulation of the alpha-2-adrenoceptors prevents IAP from inactivating the receptor system.     

Adrenergic stimulation of diacylglycerol production in genital tract smooth muscle myocytes

Summary Diacylglycerol (DAG) production has not been reported in previous studies that have characterized inositol phosphate production during alpha-1 adrenergic receptor signal transduction in the DDT₁ MF-2 genital tract myocytes. The current study sought to measure norepinephrine (NE)-stimulated DAG production in these transformed myocytes utilizing thin layer chromatography. DAG production was characterized as an alpha-1 adrenergic mediated event utilizing subtype specific adrenergic agonist and antagonists. DAG production occurred in response to physiologic concentration of NE, was apparent by 30 s and was significantly increased by 2 min. Maximal DAG production was unaffected by pretreatment of the myocytes for 96 h with testosterone, which has previously been shown to induce a doubling of alpha-1 adrenergic receptors in these cells. In contrast, testosterone pretreatment did result in a shift of the dose-response curve resulting in a significantly lower EC₅₀ for NE in the treated cells compared to control myocytes. In conclusion, these studies have confirmed that DAG production occurs as a component of alpha-1 adrenergic signal transduction in the DDT₁ MF-2 myocytes; transduction events that were modulated by testosterone resulting in increased agonist sensitivity.     

Alpha-2 adrenergic receptor subtypes indicated by [3H]yohimbine binding in human brain

Pharmacologic characterization of mammalian alpha-2 adrenergic receptors in various tissues and species has provided evidence for the existence of two alpha-2 adrenergic receptor subtypes. Prazosin and oxymetazoline have been shown to differentiate between the receptor subtypes as defined in rat tissues. In order to determine the relative proportions of these two receptor subtypes in human brain, the inhibition of the binding of the alpha-2 adrenergic antagonist [3H]yohimbine by oxymetazoline and prazosin was studied in membranes from three brain regions. Inhibition curves in membranes from the cerebral cortex and cerebellum were consistent with a single class of receptor binding sites suggesting that these two brain regions contain only one of the two subtypes. This subtype has the pharmacologic characteristics of the alpha-2A adrenergic subtype (yohimbine greater than oxymetazoline much greater than prazosin). In contrast, inhibition curves for both ligands in the human caudate nucleus were consistent with a model of two classes of binding sites in approximately equal proportions, suggesting that this tissue contains approximately equal densities of the alpha-2A and alpha-2B adrenergic receptor subtypes.     

Noradrenergic enhancement of Ca2+ responses of basal dendrites in layer 5 pyramidal neurons of the prefrontal cortex

Although the excitatory effects of noradrenaline have been thoroughly studied in the central nervous system, there is relatively little known about the adrenergic effects on Ca2+ dynamics of dendrites. In the present study, we imaged basal dendrites of layer 5 pyramidal neurons in the prefrontal cortex using two-photon microscopy. In our experiments noradrenaline, applied in the bath, enhanced excitability of layer 5 pyramidal neurons. The number of evoked action potentials following current injection to the soma increased by 44.7% on average. In the basal dendrites and spines the evoked Ca2+ responses were also markedly enhanced. Noradrenaline-induced effects could be blocked by the beta-adrenergic blocker propranolol. Our data, that activation of the noradrenergic system increases excitability of layer 5 pyramidal neurons via beta-adrenergic receptors and enhances Ca2+ signaling in basal dendrites, suggest a cellular site of action for noradrenaline to improve the integrative capabilities of dendrites.     

Absence of alpha-2 adrenergic effects on cAMP production in a genital tract smooth muscle cell line

This study sought to evaluate alpha-2 and beta adrenergic modulation of cAMP production in the DDT1 MF-2 transformed smooth muscle myocyte. After stimulation with forskolin or adrenergic agonists with or without subtype specific antagonists, cAMP production was determined. These experiments confirmed an increase of cAMP in response to forskolin, isoproterenol, epinephrine, and norepinephrine; the adrenergic stimulation was inhibited by propranolol. On the other hand, the alpha-2 agonist clonidine did not inhibit cAMP production. Likewise, alpha-2 receptor blockade did not increase cAMP production in response to epinephrine. These studies, therefore, suggest that the DDT1 MF-2 myocyte does not contain a significant population of functional alpha-2 adrenergic receptors.     

Adrenergic modulation of immune cells: an update

Sympathoadrenergic pathways are crucial to the communication between the nervous system and the immune system. The present review addresses emerging issues in the adrenergic modulation of immune cells, including: the specific pattern of adrenoceptor expression on immune cells and their role and changes upon cell differentiation and activation; the production and utilization of noradrenaline and adrenaline by immune cells themselves; the dysregulation of adrenergic immune mechanisms in disease and their potential as novel therapeutic targets. A wide array of sympathoadrenergic therapeutics is currently used for non-immune indications, and could represent an attractive source of non-conventional immunomodulating agents.     

NORADRENALINE UPTAKE INTO CEREBAL CORTEX: A HISTOCHEMICAL STUDY

Abstract— Experiments are reported which illustrate, by means of the histochemical technique of F alck and H illarp , the distribution of noradrenaline in the cerebral cortex of the rat after topical application to small areas of the pia mater. The noradrenaline is accumulated in a system of fibres and cells in the cortex by a mechanism which can be blocked by desmethylimipramine, amphetamine and phenoxybenzamine, but not by reserpine. In sections from untreated cerebral cortex the fibres contain enough noradrenaline to be visible after histochemical preparation but the cells do not. However, due to the high concentrations of noradrenaline used, the specificity of the uptake into cell bodies must remain in doubt.     

The hepatic adrenergic receptors

Summary The presence of both alpha- and beta-adrenergic receptors in liver designated the hepatic plasma membrane as a useful tool for the elucidation of the mechanisms by which the hormonal signal is transfered through the membrane via a coupling system to an amplifying entity. This process is well documented for the beta-adrenergic receptor which is linked to adenylate cyclase, whereby it modulates the cyclic AMP level. Much less is known about the alpha-adrenergic receptor.Recently, two factors have contributed to a renewed interest in alpha- and beta-adrenergic receptors in liver: i) The fact that activation of glycogenolysis in isolated liver parenchymall cells by epinephrine may be mediated by either alpha- or beta-adrenergic receptors, depending on the species or on the state of nutrition, and not only by beta-adrenergic receptors as previously thought. ii) The existence of specific adrenergic agonists and antagonists radiolabeled to a high specific activity which has permitted the characterization of adrenergic receptors in terms of nature, number, affinity and regulation.The present review will be devoted to the recent progress made in the physiological, pharmacological and biochemical characterization of alpha- and beta-adrenergic receptors in the liver.     

Possible role of somatotropic hormone in regulating the functional state of glial cells in the central nervous system

Experiments were made to study the actions of STH and its fragment capable to stimulate the growth processes on MAO activity and selective binding of catecholamines by alpha- and beta-adrenoreceptors in glial cells of the rat brain cortex. For comparison the effects of STH and its fragment on 3H-PGE1 binding were studied. STH and its fragment were found to produce no influence on MAO activity or specific binding of 3H-PGE1 with glial cells. STH and its fragment (5.10(-9) M) were found to reduce specific binding of 3H-dihydroergocryptine with beta- and alpha-adrenoreceptors of glial cells, respectively. It is suggested that STH and its fragment can modulate the function of glial cells by changing the selective action of catecholamines on subpopulations of adrenergic receptors.     

Contamination of serotonin-2 binding sites by an alpha-1 adrenergic component in assays with (3H)spiperone

(3H)Spiperone binds to two sites in mouse cortical membranes. These binding sites are discriminated by methysergide and prazosin, but not by butaclamol, lysergic acid diethylamide, or ketanserin. One of these sites is serotonergic in nature and is the authentic S-2 binding site. The other component is adrenergic and corresponds to the alpha-1 adrenoreceptor. This alpha-1 component may be present in other S-2 binding assays using (3H)spiperone, or (3H)ketanserin. No (3H)spiperone binding to dopaminergic D-2 sites was found in mouse cortex. Methods of avoiding alpha-1 contamination of S-2 binding assays are suggested.     

Effect of noradrenaline on the methaemoglobin concentration of rainbow trout red cells.

We studied the effect of noradrenaline on the methaemoglobin (metHb) concentration in rainbow trout red cells. The erythrocytes were incubated in physiological medium with or without noradrenaline and the percentage of metHb of total Hb content was measured. Noradrenaline lowered the metHb content significantly as compared to controls. To study if the effect of noradrenaline was caused by adrenergic intracellular alkalinization, cells were treated with noradrenaline + carbonic anhydrase or noradrenaline + acetazolamide. Carbonic anhydrase inhibits the adrenergic increase in intracellular pH, but did not reduce the effect of noradrenaline on the metHb concentration. Acetazolamide accentuates the increase in intracellular pH. However, there was no difference in the methaemoglobin content of noradrenaline-incubated and noradrenaline + acetazolamide-incubated cells. These results show that the effect of noradrenaline on the methaemoglobin content is independent from the adrenergic increase in intracellular pH. However, amiloride treatment inhibited the effect of noradrenaline on the methaemoglobin content, suggesting that the protein mediating sodium/proton exchange may also be involved in controlling cellular methaemoglobin levels.     

Action of dopamine on the human iris

Dopamine eye drops produce marked dilatation of the pupil in man. This mydriatic effect is inhibited by pretreatment with guanethidine. It is therefore concluded that dopamine acts indirectly via adrenergic nerve endings, rather than exerting a direct effect on adrenergic receptors in the dilator pupillae muscle. In this respect dopamine resembles the phenyl-alkylamines, such as tyramine, rather than the catecholamines, adrenaline and noradrenaline.If dopamine acts by releasing noradrenaline from adrenergic nerve endings, high concentrations of dopamine could lead to depletion of noradrenaline stores, since synthesis might be unable to keep pace with release. This could be the explantion for the orthostatic hypotension found in patients taking L-dopa for Parkinsonism.     

Alpha adrenergic stimulation reduces cyclic adenosine 3',5'-monophosphate generation in rabbit myometrium by two mechanisms

Rabbit myometrium contains postsynaptic alpha-1, alpha-2, and beta-2 adrenoreceptors. The response to endogenous catecholamines depends on the summation of interactions at these receptors and is influenced by the hormonal environment. Estrogen treatment of ovariectomized rabbits increases the alpha adrenergic contractile response whereas progesterone treatment of estrogen primed animals results in a predominance of the beta adrenergic response, which is inhibition of contractions. Of the receptor subtypes, only the alpha-2 receptor concentration is increased at physiological estrogen concentrations. However, alpha-2 receptors have not been shown to be directly involved in myometrial contraction, which appears to be mediated solely by alpha-1 adrenergic interactions. To test whether alpha-2 receptors might indirectly affect contraction by opposing interactions at the beta receptor, we examined the ability of alpha adrenergic stimulation to reduce myometrial cyclic adenosine 3',5'-monophosphate (cAMP) generation. We find that alpha-2 receptors inhibit myometrial ade adenylate cyclase through the guanine nucleotide regulatory protein, Gi. In addition, we find that activation of alpha-1 receptors also reduces cAMP generation. This interaction, which can be demonstrated in the absence but not the presence of the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine, does not appear to be mediated through Gi. These findings illustrate the complexity of adrenergic interactions in tissues containing several adrenergic subtypes.     

In vivo monitoring of evoked noradrenaline release in the rat anteroventral thalamic nucleus by continuous amperometry

Continuous amperometry coupled with untreated carbon-fibre electrodes was used in anaesthetized rats to measure the noradrenaline release evoked in the anteroventral thalamic nucleus by electrical stimulation of the dorsal noradrenergic bundle. As expected, the variations in the oxidation current detected in the anteroventral thalamic nucleus exhibited the characteristics of the in vivo noradrenaline release. They were closely correlated with stimulation and consistent with the anatomy of the noradrenergic system involved. They were abolished by the ejection of tetrodotoxin in the vicinity of the carbon-fibre electrode, diminished by clonidine, an alpha-2 agonist, and restored by yohimbine, an alpha-2 antagonist. Furthermore, the time course of these variations was dramatically increased by desipramine, a specific noradrenaline reuptake blocker. In contrast, neither dopamine nor serotonin reuptake blockers, nor the monoamine oxidase inhibitor pargyline were able to alter them. The main advantage of the present approach is its excellent time resolution. We show here for the first time that after single pulse stimulation, noradrenaline is released and eliminated in 118 milliseconds, this time lapse corresponding to the maximal period beyond which subsequent noradrenaline releases could not add up. These observations are in good agreement with the physiological relationship previously observed between impulse flow and noradrenaline overflow.     

The ontogenetic correlations of noradrenaline level and adrenergic receptor density in the rat brain

We studied the level of noradrenaline and the density of alpha 2- and beta-adrenoreceptors in the brain stem and cerebral cortex of 12-day- and 21-day-old rat fetuses, as well as of rats at the ages of 1, 2, 5, 7, 9, 16, 21, 35, and 70 days. We found a positive correlation between the level of noradrenaline in the brain stem and the density of beta-receptors in the cerebral cortex, and between the amount of alpha 2- and beta receptors in the cerebral cortex, as well as between the values of each of these indices of the neurochemical system and body weight. Significant negative correlations (r = -0.72 and r = -0.88, respectively) were found between the amount of alpha 2-adreno-receptors in the brain stem and the content of noradrenaline in this brain region, as well as in the cerebral cortex. Explanations of these positive and negative correlations between the level of noradrenaline and the amount of adrenergic receptors in the rat brain during ontogenesis are discussed.     

Reduced alpha adrenergic mediated contraction of renal preglomerular blood vessels as a function of gender and aging

As human males age, a decline in baroreflex-mediated elevation of blood pressure occurs due, at least in part, to a reduction in alpha-1 adrenergic vasoconstrictor function. Alpha adrenergic constriction is mediated by guanosine triphosphate binding Protein (G Protein) coupled signaling pathways. Alpha-1 A/C, B, and D adrenergic receptor expressions, measured by GeneChip array, are not reduced during aging in renal blood vessels of male or female rats. Alpha-1 A GeneChip expression is greater, at all ages studied, in females than in males. Prazosin binding by alpha-1 adrenergic receptors is greater in young adult female rats than in young adult male rats; however, it is reduced with aging in both male and female rats. G alpha q GeneChip expression declines while expression of adrenergic receptor kinase (GRK2) and tyrosine phosphatases (TyrP) increase with aging in male rats. The declines in alpha-1 adrenergic receptor binding and G alpha q expression and also the increases in GRK2 and TyrP expression likely relate to the age-related decline of vasoconstriction in male rats. The information that the expression of alpha-1 A adrenergic receptors is greater in female rats and (GRK2) expression does not increase during aging could relate to the gender differences in vasoconstrictor function with aging. Gene therapy to ameliorate the age-related decline in renal function could possibly reduce the need for renal dialysis. Signaling pathways such as those reviewed herein may provide an outline of the molecular pathways needed to move toward successful renal gene therapy for aging individuals.     

Comparative effects of cytochalasin D on the protein discharge induced by alpha- and beta-adrenergic or cholinergic agonists in rat exorbital lacrimal glands

Cytochalasin D altered the kinetics of peroxidase and radiolabeled protein discharge from rat exorbital lacrimal glands in vitro, in response to various secretagogues. The changes were different with each inducer. The discharge due to isoproterenol was immediately inhibited by 95%; the discharge evoked by noradrenaline via alpha-adrenergic receptors was progressively reduced and was inhibited by 50% after 30 min, whereas that evoked by carbachol was not influenced during the initial discharge period and was diminished by only 30% after 30 min. When calcium was removed from the incubation medium, the secretory responses were lowered and the inhibitory effect of cytochalasin D was still observed. The rate of protein discharge inhibition was related to the dose and was maximal with 2 X 10(-6) M cytochalasin D when the discharge resulted from cholinergic, alpha- or beta-adrenergic or dibutyryl cAMP stimulation. Cytochalasin D did not impair cellular energetics nor other stimulations induced through muscarinic or adrenergic receptors. Cytochalasin D effects could be related to interaction with actin, leading to the inhibition of the release of proteins into the incubation medium following the activation of the adrenergic system.     

Differential contribution of syntaxin 1 and SNAP-25 to secretion in noradrenergic and adrenergic chromaffin cells

We used botulinum neurotoxins (BoNT) to examine whether differences in the secretory activity of noradrenergic and adrenergic chromaffin cells are related to differences in the exocytotic machinery of these two types of bovine adrenal medulla cells. Cleavage of syntaxin and SNAP-25 by BoNT/C1 decreased in a dose-dependent way the release of both noradrenaline and adrenaline, but noradrenaline release was more sensitive to BoNT/C1. Cleavage of SNAP-25 by BoNT/A also had a larger inhibitory effect on noradrenaline release than on adrenaline release. Neither BoNT/C1 nor BoNT/A affected the intracellular Ca2+ responses induced by K+-depolarisation, and the extent of the inhibition of K+-evoked catecholamine release by selective blockers of voltage-gated Ca2+ channels was not affected by BoNT/C1. Therefore, our data do not support the hypothesis of a regulatory effect of syntaxin or SNAP-25 on the activity of Ca2+ channels. The lower sensitivity of adrenaline release to BoNT was not due to a reduced ability of the toxins to enter or to cleave their protein targets in adrenergic cells, since immunoblot analysis showed the cleavage of a larger fraction of syntaxin 1A in adrenergic cells, as compared to the cleavage in noradrenergic cells. The immunoblot analysis also showed larger amounts of syntaxin 1A in noradrenergic chromaffin cells than in adrenergic cells. Thus, in spite of a greater cleavage of syntaxin 1A in adrenergic cells by BoNT/C1, adrenaline release was less sensitive to BoNT/C1, suggesting that the release process in noradrenergic cells might be more dependent on syntaxin 1A and SNAP-25, as compared to adrenergic cells.