Presynaptic alpha-adrenoceptors in median preoptic nucleus modulate inhibitory neurotransmission from subfornical organ and organum vasculosum lamina terminalis

The median preoptic nucleus (MnPO) in the lamina terminalis receives a prominent catecholaminergic innervation from the dorsomedial and ventrolateral medulla. The present investigation used whole cell patch-clamp recordings in rat brain slice preparations to evaluate the hypothesis that presynaptic adrenoceptors could modulate GABAergic inputs to MnPO neurons. Bath applications of norepinephrine (NE; 20-50 microM) induced a prolonged and reversible suppression of inhibitory postsynaptic currents (IPSCs) and reduced paired-pulse depression evoked by stimulation in the subfornical organ and organum vasculosum lamina terminalis. These events were not correlated with any observed changes in membrane conductance arising from NE activity at postsynaptic alpha(1)- or alpha(2)-adrenoceptors. Consistent with a role for presynaptic alpha(2)-adrenoceptors, responses were selectively mimicked by an alpha(2)-adrenoceptor agonist (UK-14304) and blockable with an alpha(2)-adrenoceptor antagonist (idazoxan). Although the alpha(1)-adrenoceptor agonist cirazoline and the alpha(1)-adrenoceptor antagonist prazosin were without effect on these evoked IPSCs, NE was noted to increase (via alpha(1)-adrenoceptors) or decrease (via alpha(2)-adrenoceptors) the frequency of spontaneous and tetrodotoxin-resistant miniature IPSCs. Collectively, these observations imply that both presynaptic and postsynaptic alpha(1)- and alpha(2)-adrenoceptors in MnPO are capable of selective modulation of rapid GABA(A) receptor-mediated inhibitory synaptic transmission along the lamina terminalis and therefore likely to exert a prominent influence in regulating cell excitability within the MnPO.     

Estrogen receptor alpha rapidly activates the IGF-1 receptor pathway

Estrogen and insulin-like-growth factor 1 (IGF-1) are potent mitogenic stimuli that share important properties in the control of cellular proliferation. However, the coupling between the signaling cascades of estrogen receptors alpha and beta and the IGF-1 receptor (IGF-1R) is poorly understood. Therefore, we selectively transfected estrogen receptor alpha or beta in COS7 and HEK293 cells, which contain IGF-1R. In presence of estrogen receptor alpha but not beta, 17beta-estradiol (E2) rapidly induces phosphorylation of the IGF-1R and the extracellular signal-regulated kinases 1/2. Furthermore, upon stimulation with E2, estrogen receptor alpha but not beta bound rapidly to the IGF-1R in COS7 as well as L6 cells, which express all investigated receptors endogenously. Control experiments in the IGF-1R-deficient fibroblast cell line R(-) showed that after stimulation with E2 only estrogen receptor alpha bound to the transfected IGF-1R. Overexpression of dominant negative mitogen-activated protein kinases kinase inhibited this effect. Finally, estrogen receptor alpha but not beta is required to induce the activation of the estrogen receptor-responsive reporter ERE-LUC in IGF-1-stimulated cells. Taken together, these data demonstrate that ligand bound estrogen receptor alpha is required for rapid activation of the IGF-1R signaling cascade.     

Norepinephrine induces apoptosis in skin melanophores by attenuating cAMP-PKA signals via alpha2-adrenoceptors in the medaka, Oryzias latipes

The density of skin melanophores in many teleost fish decreases during long-term adaptation to a white background. Using the medaka, Oryzias latipes, we previously reported that apoptosis is responsible for the decrease in melanophores, and that a sympathetic neurotransmitter, norepinephrine (NE), induces their apoptosis in skin tissue cultures. In this study, we show that NE-induced apoptosis of melanophores is mediated by the activation of alpha2-adrenoceptors. Clonidine, an alpha2-adrenoceptor agonist, induced apoptotic melanophore death in skin organ culture, while phenylephrine, an alpha1-adrenoceptor agonist, had no effect. NE-induced apoptosis was diminished by an alpha2-adrenoceptor antagonist, yohimbine, but an alpha1-adrenoceptor antagonist, prazosin, did not abrogate the effect of NE. Furthermore, forskolin inhibited NE-induced apoptosis, while an inhibitor of PKA, H-89, mimicked the effect of NE. These results suggest that NE induces apoptosis in melanophores by attenuating cAMP-PKA signaling via alpha2-adrenoceptors.     

Visualizing activation of opioid circuits by internalization of G protein-coupled receptors

Mu-opioid receptor (MOR) and opioid receptor-like receptor (ORL-1) circuits in the limbic hypothalamic system are important for the regulation of sexual receptivity in the female rat. Sexual receptivity is tightly regulated by the sequential release of estrogen and progesterone from the ovary suggesting ovarian steroids regulate the activity of these neuropeptide systems. Both MOR and ORL-1 distributions overlap with the distribution of estrogen and progesterone receptors in the hypothalamus and limbic system providing a morphological substrate for interaction between steroids and the opioid circuits in the brain. Both MOR and ORL-1 are receptors that respond to activation by endogenous ligands with internalization into early endosomes. This internalization is part of the mechanism of receptor desensitization or down regulation. Although receptor activation and internalization are separate events, internalization can be used as a temporal measure of circuit activation by endogenous ligands. This review focuses on the estrogen and progesterone regulation of MOR and ORL-1 circuits in the medial preoptic nucleus and ventromedial nucleus of the hypothalamus that are central to modulating sexual receptivity.     

Effects of temperature on alpha adrenoceptors in limb veins: role of receptor reserve

In cutaneous veins of the dog, cooling augments the response to sympathetic nerve stimulation and exogenous norepinephrine (NE). The postjunctional alpha adrenoceptors in this blood vessel belong to both the alpha 1 and alpha 2 subtypes. Cooling augments alpha 2-adrenergic responses (presumably because of an increased receptor affinity), but depresses alpha 1-adrenergic responses (presumably because of a direct inhibitory effect on the contractile process). When agonists of high efficacy such as NE or phenylephrine are used, an alpha 1-adrenoceptor reserve is present that buffers the response from the inhibitory effect of cooling. This allows the potentiating effect of cold on the alpha 2-adrenergic component of the response to catecholamines to predominate, and the contractile response to exogenous NE and sympathetic nerve stimulation is augmented. By contrast, in deep veins of the limb, cold reduces the contractions evoked by alpha 1- and alpha 2-adrenergic activation. This can be explained best by the absence of a receptor reserve for alpha 1-adrenergic agonists of high efficacy, combined with a reduced density of postjunctional alpha 2 adrenoceptors.     

Expression and function of alpha-adrenoceptors in zebrafish: drug effects, mRNA and receptor distributions

The alpha2-adrenoceptors are G-protein-coupled receptors that mediate many of the physiological effects of norepinephrine and epinephrine. Mammals have three subtypes of alpha2-adrenoceptors, alpha2A, alpha2B and alpha2C. Zebrafish, a teleost fish used widely as a model organism, has five distinct alpha2-adrenoceptor genes. The zebrafish has emerged as a powerful tool to study development and genetics, with many mutations causing diseases reminiscent of human diseases. Three of the zebrafish adra2 genes code for orthologues of the mammalian alpha2-adrenoceptors, while two genes code for alpha2Da- and alpha2Db- adrenoceptors, representing a duplicated, fourth alpha2-adrenoceptor subtype. The three different mammalian alpha2-adrenoceptor subtypes have distinct expression patterns in different organs and tissues, and mediate different physiological functions. The zebrafish alpha2-adrenergic system, with five different alpha2-adrenoceptors, appears more complicated. In order to deduce the physiological functions of the zebrafish alpha2-adrenoceptors, we localized the expression of the five different alpha2-adrenoceptor subtypes using RT-PCR, mRNA in situ hybridization, and receptor autoradiography using the radiolabelled alpha2-adrenoceptor antagonist [ethyl-3H]RS-79948-197. Localization of the alpha2A-, alpha2B- and alpha2C-adrenoceptors in zebrafish shows marked conservation when compared with mammals. The zebrafish alpha2A, alpha2Da, and alpha2Db each partially follow the distribution pattern of the mammalian alpha2A: a possible indication of subfunction partitioning between these subtypes. The alpha2-adrenergic system is functional in zebrafish also in vivo, as demonstrated by marked locomotor inhibition, similarly to mammals, and lightening of skin colour induced by the specific alpha2-adrenoceptor agonist, dexmedetomidine. Both effects were antagonized by the specific alpha2-adrenoceptor antagonist atipamezole.     

Influence of development and reduction of fat stores on the antilipolytic alpha 2-adrenoceptor in hamster adipocytes: comparison with adenosine and beta-adrenergic lipolytic responses

The response of the hamster adipocyte to various lipolytic (beta-adrenergic) and antilipolytic (alpha(2)-adrenergic and adenosine-dependent) stimuli was studied during the development and after cold-induced regression of fat stores. Alpha(2)-adrenergic binding ([(3)H]clonidine binding sites) was also investigated. Adipocytes came from young animals (4-5 weeks), adults (20-25 weeks), and adults submitted to a 6-week cold exposure (6 degrees C) that promoted a large decrease in fat stores and in fat cell size. The lipolytic response induced by isoproterenol (beta-agonist) was equivalent in the different groups. Adenosine and alpha(2)-adrenergic antilipolytic effects were estimated through the inhibition of theophylline-induced lipolysis by phenylisopropyladenosine and clonidine, respectively. The adenosine effect was unchanged in all the groups. In contrast, the alpha(2)-adrenergic effect, which was not present in young hamsters, increased simultaneously with fat cell size, was fully effective in adult hamsters, and had completely disappeared in small adipocytes from cold-exposed hamsters. In fat cell ghosts, alpha(2)-adrenoceptors ([(3)H]clonidine binding sites), followed similar modifications: they increased with fat cell enlargement and disappeared after cell size reduction following cold exposure. These results suggest that: 1) the increased alpha(2)-adrenergic antilipolytic response which is concomitant with fat cell enlargement could partly explain the growth-related decrease in the previously reported lipolytic effect of epinephrine; 2) the alpha(2)-receptivity of the adipocyte seems to be strictly fat cell size-dependent while the beta-adrenergic and adenosine responses are unaffected; and 3) the regulation in the adipocytes of the adenosine, alpha(2)- and beta-receptors seems to be unrelated.-Carpene, C., M. Berlan, and M. Lafontan. Influence of development and reduction of fat stores on the antilipolytic alpha(2)-adrenoceptor in hamster adipocytes: comparison with adenosine and beta-adrenergic lipolytic responses.     

Haloperidol-mediated phosphoinositide hydrolysis via direct activation of alpha1-adrenoceptors in frontal cerebral rat cortex.

In addition to its effect on D2 dopamine receptor blockades, haloperidol is able to interact with multiple neurotransmitters (NTs). Its action on phosphoinositide (PI) turnover was studied on cerebral cortex preparations. It induces an increase in inositol phosphate (IP) accumulation, which was only blunted by the alpha1-adrenoceptor blocker prazosin. Haloperidol maximal effect (Emax) was less than the effect of the full agonist norepinephrine (NE), and dose-response curves for both NE in the presence of submaximal doses of haloperidol and haloperidol in the presence of Emax doses of NE showed that haloperidol behaves as a partial agonist of cerebral alpha1-adrenoceptors. Its effect on PI hydrolysis is mediated through phospholipase C activation, as 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate (NCDC) and 1-[6-([(17beta)-3-methoxyestra- 1,3,5(10)-trien-17-yl]amino)hexyl]-1H-pyrrole-2,5-dione) (U-73122) were able to abrogate both haloperidol and NE actions. Further, the typical neuroleptic exerts a direct activation of alpha1-adrenoceptors as its actions were not modified by cocaine and persisted in spite of chemical rat cerebral denervation with 6-hydroxydopamine (6-OHDA). The possibility that this agonistic action on alpha1-adrenoceptors would be involved in haloperidol side effects is also discussed.     

Involvement of norepinephrine activity in the regulation of alpha 1 adrenergic receptors in the medial preoptic nucleus of estradiol-treated rats

To establish whether the diurnal decrease in the density of alpha 1 receptors observed in the medial preoptic nucleus (MPN) of estrogen (E2)-treated rats is related to the concomitant diurnal increase in norepinephrine (NE) turnover rates, we quantitated the density of [3H]-Prazosin binding to alpha 1 receptors after blockade of NE turnover with alpha-methyl-paratyrosine (alpha MPT). A series of preliminary studies was performed to rule out an interference of this drug with [3H]-Prazosin binding to alpha 1 adrenergic receptors in vitro and in vivo. Incubation of brain slices with alpha MPT produced a dose-dependent inhibition of [3H]-Prazosin binding to alpha 1 adrenergic receptors with an IC50 of approximately 6 mM. Scatchard analysis demonstrated that alpha MPT exhibited a simple competitive interaction with [3H]-Prazosin binding sites as shown by an increase in the apparent dissociation constant (Kd) of the ligand and no change in the number of alpha 1 receptors (Bmax). In contrast, preincubation of brain slices with alpha MPT and prior in vivo administration of alpha MPT did not affect [3H]-Prazosin binding to alpha 1 adrenergic receptors. Once we established that alpha MPT could be used to suppress NE turnover without interfering with the measurement of alpha 1 receptor densities, we repeatedly injected this drug to ovariectomized (OVX) and E2-implanted rats. The density of alpha 1 adrenergic receptors in MPN was quantitated autoradiographically. Blockade of NE turnover with alpha MPT only partially prevented the reduction in alpha 1 receptor density observed in the E2-treated rats, suggesting that the decrease in the level of [3H]-Prazosin binding sites cannot be completely ascribed to increased NE turnover rates.     

Pharmacological characterization of alpha1- and beta-adrenergic synergism of 5'DII activity in rat brown adipocytes

The role of adrenoceptor subtypes was studied in rat brown adipose tissue (BAT). The type II 5'-deiodinase (5'DII) was activated in response to simultaneous stimulation by beta3- and alpha1-adrenergic agonists, BRL 37344 or CGP 12177, and cirazoline, in brown adipocytes. Inhibition of the alpha1- and beta-adrenergic phenylephrine-stimulated 5'DII activity was obtained by the alpha1-adrenergic antagonists in the order of prazosin >/= wb 4101 > 5-methylurapidil. In comparison, the binding of [3H]prazosin to rat BAT plasma membranes was inhibited by alpha1-adrenergic antagonists in the order of prazosin > WB 4101 = benoxathian > 5-methylurapidil. Although the order of the alpha1-adrenergic competition seemed to be rather typical for the alpha1B-adrenergic receptors, a molecular analysis on adrenoceptor mRNAs should be made to confirm the exact alpha1-adrenergic subtypes at the level of brown adipocytes, since the possibility of a mixture of different receptor subtypes in brown fat cells and/or tissue may interact with the pharmacological characterization. Thus, specific alpha1- and beta-adrenoceptor subtypes participate in the regulation of 5'DII activity in the rat brown adipocytes, and therefore, an impaired alpha1- and beta-adrenergic co-work may be involved in a defective BAT function, e.g., in obese Zucker rats, too. An interesting possibility is that the decreased number of alpha1-adrenoceptors in the BAT of obese Zucker rats is due to the decrease in the alpha1B-adrenoceptor subtype which would further be involved especially in the regulation of BAT 5'DII activity.     

beta-Adrenergic stimulated lipolysis in pony adipocytes is exclusively via a beta2-subtype and is not affected by lactation

Catecholamines are important lipolytic agents in horses and ponies but the nature of the adrenergic receptor subtype distribution in their adipocytes is uncertain. A first objective was to identify the beta-adrenergic receptor subtype(s) present in adipocytes from horses and ponies. A second objective was to evaluate if the lipolytic responsiveness of isolated adipocytes to beta-adrenergic agonists is altered during lactation, a condition known to affect markedly maternal fat metabolism. Isoproterenol and salbutamol elicited strong lipolytic responses in adipocytes isolated from horse and pony subcutaneous adipose tissue. There were weak lipolytic responses to norepinephrine, dobutamine and BRL37344. The weak lipolytic response to NE compared to isoproterenol or salbutamol suggests an antilipolytic action from alpha2-adrenergic receptors. The relative order of potency for the beta-adrenergic agonists was isoproterenol>/=salbutamol>dobutamine=BRL37344. There was expression of beta2-adrenergic receptor mRNA in pony and horse adipose tissues, as estimated by relative RT-PCR, but no expression of mRNAs for beta1- or beta3-adrenergic receptors. Early lactation did not alter the lipolytic responses to beta-adrenergic agonists, nor the expression of beta2-adrenergic receptor mRNA. Thus, these results indicate a dominant if not exclusive presence of beta2-adrenergic receptors in pony and horse adipocytes that is not affected by lactation.     

Conjugated estradiol increases female sexual receptivity in response to oxytocin infused into the medial preoptic area and medial basal hypothalamus

The ovarian steroid estradiol (E) has been found to increase both receptor affinity and release of the neuropeptide oxytocin (OT) in plasma membrane preparations. Therefore, we hypothesized that E conjugated to bovine serum albumin at position 6 (E-6-BSA) would increase behavioral responsiveness to OT. Preliminary results showed that 200 ng/microl of E-6-BSA increased sexual receptivity slightly, but not significantly. Therefore, this dose was used as a subthreshold dose to test whether it would increase sexual responsiveness when infused in combination with 100 ng/microl OT. After recovery from cannula implantation surgery animals were injected with 0.5 microg E benzoate daily for 3 days before testing. On the fourth day, after a baseline preinfusion test rats were infused bilaterally with E-6-BSA alone or with OT, OT with BSA, or conjugated progesterone, luteinizing hormone-releasing hormone equimolar to OT alone, or with E-6-BSA or conjugated progesterone alone. When infused into either the medial preoptic area-anterior hypothalamus or the medial basal hypothalamus the combination of OT and E-6-BSA significantly increased sexual receptivity over receptivity after artificial cerebrospinal fluid control infusions. Neither bilateral infusions of OT in combination with conjugated progesterone nor E-6-BSA in combination with luteinizing hormone-releasing hormone enhanced sexual receptivity. Results presented here strongly support the conclusion that some of the effects that E has in sensitizing brain systems to the facilitating effects of OT occur at the membrane level in the medial preoptic area-anterior hypothalamus and medial basal hypothalamus.     

Phorbol ester effects on alpha 1-adrenoceptor binding and phosphatidylinositol metabolism in cultured vascular smooth muscle cells

Tumor promoting phorbol esters stimulate Ca++ phospholipid-dependent protein kinase C. It has been suggested that this enzyme regulates the functional properties of different cell membrane receptors. In this study we investigated the effect of phorbol esters on alpha 1-adrenoceptor binding and phosphatidylinositol metabolism in cultured smooth muscle cells derived from rabbit aorta. Treatment of these cells with biologically active phorbol esters for 15 min. to 2 hours caused a marked decrease of norepinephrine stimulation of inositol phospholipid metabolism and a 3 fold decrease in agonist affinity for 125I-HEAT binding to alpha 1-adrenoceptors in the intact smooth muscle cells. The ability of phorbol esters to modulate alpha 1-adrenoceptor responsiveness suggests that activation of protein kinase C may represent an important mechanism regulating alpha 1-adrenergic receptor functional properties.     

Functional alpha 2-adrenoceptors in rat adipocytes: inhibition of cyclic AMP accumulation

Alpha 2-adrenoceptor activation inhibits cyclic AMP accumulation in fat cells from many species. However, the presence of alpha 2-adrenoceptors in rat adipocytes has been difficult to demonstrate. We observed that alpha 2-adrenergic activation inhibits forskolin-stimulated cyclic AMP accumulation both in rat and hamster adipocytes; UK 14304, p-amino clonidine and clonidine were the agents with higher efficacy. The effect of UK 14304 was blocked by yohimbine but not by prazosin demonstrating the involvement of alpha 2-adrenoceptors. Pertussis toxin blocked the alpha 2-adrenergic effect. Our results demonstrate the presence in rat fat cells of alpha 2-adrenoceptors coupled to adenylate cyclase via "Gi".     

Brain adrenoreceptors in rats with inherited arterial hypertension due to emotional stress

For the study of genetic and physiological mechanisms of inherited stress-sensitive arterial hypertension, specific binding of ligands of alpha 1-, alpha 2- and beta-adrenoceptors was measured in 2 strains of rats: Wistar normotensive and ISSAH rats (rats with inherited stress-sensitive arterial hypertension). The maximal binding sites (Bmax) and apparent dissociation constants (Kd) were studied with the alpha 1-adrenergic antagonist 3H-prazosin, alpha 2-adrenergic agonist 3H-clonidine and 3H-dihydroalprenolol, a beta 1-receptor antagonist. Four brain regions were investigated: frontal cortex, hypothalamus, pons and medulla oblongata. In comparison with normotensive controls, hypertensive rats had significantly greater density of the alpha 1-adrenoceptors in the medulla oblongata. However, the number of hypothalamic alpha 1-adrenoceptors was significantly reduced in these animals. The same significantly lower alpha 2-adrenoreceptor density was found in the hypothalamus and the pons, and lower, beta-adrenoceptors density in the medulla oblongata. It was concluded that brain adrenoceptors are involved in the mechanisms of development of inherited stress-sensitive hypertensive syndrome.     

Agonist-dependent phosphorylation of the alpha 2-adrenergic receptor by the beta-adrenergic receptor kinase

Desensitization of the beta-adrenergic receptor, a receptor which is coupled to the stimulation of adenylate cyclase, may be regulated via phosphorylation by a unique protein kinase. This recently discovered enzyme, known as the beta-adrenergic receptor kinase, only phosphorylates the agonist-occupied form of the beta-adrenergic receptor. To assess whether receptors coupled to the inhibition of adenylate cyclase might also be substrates, we examined the effects of beta-adrenergic receptor kinase on the partially purified human platelet alpha 2-adrenergic receptor. Phosphorylation of the reconstituted alpha 2-adrenergic receptor was dependent on agonist occupancy and was completely blocked by coincubation with alpha 2-antagonists. The time course of phosphorylation of the alpha 2-adrenergic receptor was virtually identical to that observed with the beta-adrenergic receptor with maximum stoichiometries of 7-8 mol of phosphate/mol of receptor in each case. In contrast, the alpha 1-adrenergic receptor, which is coupled to stimulation of phosphatidylinositol hydrolysis, is not a substrate for the beta-adrenergic receptor kinase. These results suggest that receptors coupled to either stimulation or inhibition of adenylate cyclase may be regulated by an agonist-dependent phosphorylation mediated by the beta-adrenergic receptor kinase.     

Differential effect of pertussis toxin on the affinity state for agonists of renal alpha 1- and alpha 2- adrenoceptors

The effect of pertussis toxin treatment on the guanine nucleotide-induced modulation of the affinity of renal alpha 1- and alpha 2-adrenergic receptors was investigated. Pretreatment of rats with pertussis toxin did not induce any change in the number of or affinity for antagonists of alpha 1- or alpha 2-receptors studied using [3H]prazosin and [3H]yohimbine, respectively. Guanyl-5'-yl imidodiphosphate induced an "up-shift" in the number of alpha 2-adrenergic receptors; this up-shift was not observed for alpha 1-adrenergic receptors. Pertussis toxin treatment decreased the affinity of epinephrine for the [3H]yohimbine-binding sites and reduced the ability of guanine nucleotides to modulate alpha 2-adrenoceptor agonist affinity. The regulation by guanine nucleotides of alpha 1-adrenoceptor affinity for agonists was not altered. These results suggest that the modulation of alpha 1- and alpha 2-adrenoceptors by guanine nucleotides is probably exerted through different molecular entities.     

Late expression of alpha 2-adrenergic-mediated antilipolysis during differentiation of hamster preadipocytes.

In order to study the ontogenesis of the beta- and alpha 2-adrenergic control of lipolysis during the adipose conversion process, a model based on preadipocytes isolated from the stromal-vascular fraction of hamster adipose tissue was developed. When cultured in an ITT (insulin, transferrin, triiodothyronine) medium supplemented with 2% fetal calf serum, adipose precursors differentiated into adipose-like cells. On 8-day-post-confluent differentiating preadipocytes, the rank order of potency of activation of lipolysis by various beta-adrenergic agonists (BRL37344 greater than norepinephrine = isoproterenol greater than epinephrine greater than fenoterol) was equivalent to that determined in mature adipocytes isolated from adult hamster adipose tissue. On 8-day-post-confluent differentiating preadipocytes, phenylisopropyladenosine (A1-adenosine agonist) and prostaglandin E1 evoked a strong antilipolytic response whereas that evoked by UK 14304 and clonidine (alpha 2-adrenergic agonists) remained undetectable at this step of differentiation. The activity of UK 14304 and clonidine only appeared on 20- to 25-day-post-confluent differentiating preadipocytes. They induced dose-dependent antilipolysis with a maximal effect reaching 80-85% inhibition of adenosine deaminase-stimulated lipolysis. Their action was blocked by increased concentrations of different alpha 2-adrenergic antagonists with the following order of potency, RX 821002 greater than phentolamine much greater than yohimbine. This order of potency was similar to that determined on mature adipocytes isolated from adult hamsters. Both the density of the alpha 2-adrenoceptors, identified with the selective alpha 2-adrenergic radioligand [3H]RX-821002 (19 +/- 1 vs. 30 +/- 1 fmol/mg protein: P less than 0.01) and the amount of Gi proteins identified by pertussis toxin-catalyzed ADP-ribosylation (31 +/- 4 vs. 43 +/- 4% of the amount defined in mature fat cells from adult hamsters: P less than 0.05) were significantly increased between 8 days and 20-25 days after confluence, explaining the late emergence of the alpha 2-adrenergic control of lipolysis during preadipocyte differentiation. In conclusion, the late emergence of the alpha 2-adrenergic control of lipolysis, which is also supported by previous data obtained in vivo that demonstrated the age and/or the fat cell size dependence of alpha 2-adrenoreceptor expression in mature adipocytes, allows the alpha 2-adrenoceptor to be considered as a marker of adipocyte hypertrophy.     

Hypothalamic adrenergic receptor changes in the metabolic syndrome of genetically obese (ob/ob) mice

The genetically, seasonally, and diet-induced obese, glucose-intolerant states in rodents, including ob/ob mice, have each been associated with elevated hypothalamic levels of norepinephrine (NE). With the use of quantitative autoradiography on brain slices of 6-wk-old obese (ob/ob) and lean mice, the adrenergic receptor populations in several hypothalamic nuclei were examined. The binding of [(125)I]iodocyanopindolol to beta(1)- and beta(2)-adrenergic receptors in ob/ob mice was significantly increased in the paraventricular hypothalamic nucleus (PVN) by 30 and 38%, in the ventromedial hypothalamus (VMH) by 23 and 72%, and in the lateral hypothalamus (LH) by 10 and 15%, respectively, relative to lean controls. The binding of [(125)I]iodo-4-hydroxyphenyl-ethyl-aminomethyl-tetralone to alpha(1)-adrenergic receptors was also significantly increased in the PVN (26%), VMH (67%), and LH (21%) of ob/ob mice. In contrast, the binding of [(125)I]paraiodoclonidine to alpha(2)-adrenergic receptors in ob/ob mice was significantly decreased in the VMH (38%) and the dorsomedial hypothalamus (17%) relative to lean controls. This decrease was evident in the alpha(2A)- but not the alpha(2BC)-receptor subtype. Scatchard analysis confirmed this decreased density of alpha(2)-receptors in ob/ob mice. Together with earlier studies, these changes in hypothalamic adrenergic receptors support a role for increased hypothalamic NE activity in the development of the metabolic syndrome of ob/ob mice.