Blockade of sympathetic ganglia improves vascular dysfunction in septic shock.

Sepsis/septic shock activates the sympathetic nervous system (SNS) to deal with the infection stress. However, an imbalanced or maladaptive response due to excessive or uncontrolled activation characterizes autonomic dysfunction. Our hypothesis was that reducing this excessive activation of the autonomic nervous system would impact positively in sepsis. Using ganglionic blockers as a pharmacological approach, the main aim of the present report was to assess the role of ganglionic transmission in the vascular dysfunction associated with sepsis.Sepsis was induced in rats by cecal ligation and puncture (CLP). One hour after CLP surgery, rats were treated subcutaneously with hexamethonium (15 mg/kg; ganglionic blocker), pentolinium (5 mg/kg; a blocker with a higher selectivity for sympathetic ganglia compared to hexamethonium), or vehicle (PBS). Basal blood pressure and the response to adrenergic agonists were evaluated at 6 and 24 h after CLP surgery. Reactivity to vasoconstrictors, nitric oxide (NO) synthase 2 (NOS-2) expression, IL-1 and TNF plasma levels, and density of α1 adrenergic receptors were evaluated in the aorta 24 h after CLP.Septic shock resulted in hypotension and hyporesponsiveness to norepinephrine and phenylephrine, increased plasma cytokine levels and NOS-2 expression in the aorta, and decreased α1 receptor density in the same vessel. Pentolinium but not hexamethonium recovered responsiveness and α1 adrenergic receptor density in the aorta. Both blockers normalized the in vivo response to vasoconstrictors, and reduced plasma IL-1 and NOx levels and NOS-2 expression in the aorta.Blockade of ganglionic sympathetic transmission reduced the vascular dysfunction in experimental sepsis. This beneficial effect seems to be, at least in part, due to the preservation of α1 adrenergic receptor density and to reduced NOS-2 expression and may lead to adjuvant ways to treat human sepsis.

Interplay between nitric oxide and gonadotrophin-releasing hormone in the neuromodulation of the corpus luteum during late pregnancy in the rat.

BACKGROUND: Nitric oxide and GnRH are biological factors that participate in the regulation of reproductive functions. To our knowledge, there are no studies that link NO and GnRH in the sympathetic ganglia. Thus, the aim of the present work was to investigate the influence of NO on GnRH release from the coeliac ganglion and its effect on luteal regression at the end of pregnancy in the rat. METHODS: The ex vivo system composed by the coeliac ganglion, the superior ovarian nerve, and the ovary of rats on day 21 of pregnancy was incubated for 180 min with the addition, into the ganglionic compartment, of L-NG-nitro arginine methyl ester (L-NAME), a non-selective NO synthase inhibitor. The control group consisted in untreated organ systems. RESULTS: The addition of L-NAME in the coeliac ganglion compartment decreased NO as well as GnRH release from the coeliac ganglion. In the ovarian compartment, and with respect to the control group, we observed a reduced release of GnRH, NO, and noradrenaline, but an increased production of progesterone, estradiol, and expression of their limiting biosynthetic enzymes, 3ß-HSD and P450 aromatase, respectively. The inhibition of NO production by L-NAME in the coeliac ganglion compartment also reduced luteal apoptosis, lipid peroxidation, and nitrotyrosine, whereas it increased the total antioxidant capacity within the corpora lutea. CONCLUSION: Collectively, the results indicate that NO production by the coeliac ganglion modulates the physiology of the ovary and luteal regression during late pregnancy in rats.

Long-term potentiation is differentially expressed in rostral and caudal neurons in the superior cervical ganglion of normal and hypertensive rats.

Neurons in the superior cervical ganglia (SCG) are classified as rostral and caudal according to their regional locations. Although diverse phenotypes have been reported for these two subpopulations, differences in neuroplasticity, like long-term potentiation (LTP), have not been characterized. Here, we explored possible regional differences of LTP expression in rostral and caudal neurons of the SCG in control rats, Wistar and Wistar Kyoto (WKy), and in the spontaneously hypertensive rats (SHR) as a model of hypertension. We characterized the expression of gLTP evoked by a tetanic train (40 Hz, 3 s) in an in vitro SCG preparation. gLTP was recorded in rostral and caudal neurons at 8-weeks-old (wo) in Wistar rats, 6-wo and 12-wo in SHR and WKy rats. We found that gLTP was differentially expressed; gLTP was larger in caudal neurons in Wistar and adult WKy rats. In adult 12-wo hypertensive SHR, gLTP was expressed in caudal but not in rostral neurons. In contrast, in 6-wo pre-hypertensive SHR, gLTP was expressed in rostral but not in caudal neurons; while in 6-wo WKy, gLTP was expressed in caudal but not in rostral neurons. The lack of gLTP expression in caudal neurons of 6-wo SHR was not due to a GABAergic modulation because several GABA-A receptor antagonists failed to unmask gLTP. Data show that neuroplasticity, particularly gLTP expression, varied according to the ganglionic region. We propose that differential regional expression of gLTP may be correlated with selective innervation on different target organs.

Ganglionic Long-Term Potentiation in Prehypertensive and Hypertensive Stages of Spontaneously Hypertensive Rats Depends on GABA Modulation.

The sympathetic nervous system (SNS) regulates body functions in normal and pathological conditions and is characterized by the presence of a neuroplastic phenomenon, termed ganglionic long-term potentiation (gLTP). In hypertension, either in spontaneously hypertensive rats (SHR) or in humans, sympathetic hyperfunction, such as elevated SNS outflow and changes in synaptic plasticity have been described. Because enhanced SNS outflow is detected in the hypertensive stage and, more importantly, in the prehypertensive phase of SHR, here we explored whether synaptic plasticity, particularly gLTP, was modified in the superior cervical ganglia (SCG) of prehypertensive SHR. Furthermore, considering that GABA modulates sympathetic synaptic transmission and gLTP in Wistar rats, we studied whether GABA might modulate gLTP expression in SHR. We characterized gLTP in the SCG of young prehypertensive 6-week-old (wo) and adult hypertensive (12 wo) SHR and in the SCG of Wistar Kyoto (WKy) normotensive control rats of the same ages. We found that gLTP was expressed in 6 wo SHR, but not in 12 wo rats. By contrast, in WKy, gLTP was expressed in 12 wo, but not in 6 wo rats. We also found that gLTP depends on GABA modulation, as blockade of GABA-A subtype receptors with its antagonist bicuculline unmasked gLTP expression in adult SHR and young WKy. We propose that (1) activity-dependent changes in synaptic efficacy are altered not only during hypertension but also before its onset and (2) GABA may play a modulatory role in the changes in synaptic plasticity in SHR, because the blockade of GABA-A receptors unmasked the expression of gLTP. These early changes in neuroplasticity and GABA modulation of gLTP could be part of the sympathetic hyperfunction observed in hypertension.

Prolactin modulates luteal regression from the coeliac ganglion via the superior ovarian nerve in the late-pregnant rat.

There is considerable evidence of the neuroendocrine control involved in luteal regression in the rat. In addition, circulating prolactin (PRL), which increases during the night before parturition, may gain access to the coeliac ganglion (CG), indirectly impacting the physiology of the ovary because of the known connection between the CG and the ovary via the superior ovarian nerve (SON). In this work we investigated in the CG-SON-ovary system and whether PRL added to the CG has an impact, indirectly via the SON, on luteal regression on Day 21 of pregnancy. The system was incubated without (control) or with PRL added to the CG. We measured the ovarian release of progesterone (P), oestradiol and prostaglandin F2 alpha (PGF2α) by radioimmunoassay, and nitrites (NO) by the Griess method. Luteal mRNA expression of 3ß-hydroxysteroid dehydrogenase (3ß-HSD), 20α-HSD, aromatase, inducible nitric oxide synthase (iNOS) and apoptosis regulatory factors was analysed by reverse transcription-polymerase chain reaction. P release, the expression of Bcl-2 and the Bcl-2:Bax ratio was lower than control preparations, while the expression of 20α-HSD and the release of NO and PGF2α were higher in the experimental group. In conclusion, PRL acts at the CG and, by a neural pathway, modulates luteal function at the end of pregnancy.

Neuromodulation of the luteal regression: presence of progesterone receptors in coeliac ganglion.

NEW FINDINGS: What is the central question of this study? The processes involved in luteal involution have not yet been clarified and, in general, have been studied only from a hormonal point of view. We investigated whether progesterone, from the coeliac ganglion through the superior ovarian nerve, is able to modify the luteal regression of late pregnancy in the rat. What is the main finding and its importance? We showed that the luteal regression might be reversed by the neural effect of progesterone and demonstrated the presence of its receptors in the coeliac ganglion. This suggests that the peripheral neural pathway, through neuron-hormone interaction, represents an additional mechanism to control luteal function in addition to the classical endocrine regulation. The corpus luteum (CL) is a transitory endocrine gland that produces progesterone (P). At the end of its useful life, it suffers a process of functional and structural regression until its complete disappearance from the ovary. To investigate whether P is able to regulate the process of luteal regression through the peripheral neural pathway, we used the coeliac ganglion (CG)-superior ovarian nerve-ovary system from rats on day 21 of pregnancy. We stimulated the CG with P and analysed the functional regression through ovarian P release measured by radioimmunoassay, expression by RT-PCR and activity of luteal 3ß- and 20α-hydroxysteroid dehydrogenase (anabolic and catabolic P enzymes, respectively). The luteal structural regression was evaluated through a study of apoptosis measured by TUNEL assay and the expression of apoptotic factors, such as Bcl-2, Bax, Fas and Fas ligand (FasL) by RT-PCR. To explore whether the effects mediated by P on the CL may be associated with P receptors, their presence in the CG was investigated by immunohistochemistry. In the group stimulated with P in the CG, the ovarian P release and the 3ß-hydroxysteroid dehydrogenase activity increased, whereas the expression and activity of 20α-hydroxysteroid dehydrogenase decreased. In addition, a decrease in the number of apoptotic nuclei and a decrease of the expression of FasL were observed. We demonstrated the presence of P receptors in the CG. Overall, our results suggest that the regression of the CL of late pregnancy may be reprogrammed through the peripheral neural pathway, and this effect might be mediated by P bound to its receptor in the CG.

An increase in glucose concentration in the lateral ventricles of the brain induces changes in autonomic nervous system activity.

OBJECTIVE: Changes in glucose levels mobilize a neuroendocrine response that prevents or corrects glycemia. The hypothalamus is the main area of the brain that regulates glycemic homeostasis. Metabolic diseases, such as obesity and diabetes, are related to imbalance of this control. The modulation of autonomic nervous system (ANS) activity is mediated by neuronal hypothalamic pathways. In the present work, we investigate whether glucose concentration in the hypothalamic area changes ANS activity. METHODS: Glucose was administered intracerebroventricularly to 90-day-old rats, and samples of blood were collected during brain glucose infusion to measure the blood glucose and insulin levels. The electric activity of the superior vagus nerve and superior sympathetic ganglion was directly registered. RESULTS: Glucose 5·6 mM infused in the hypothalamus induced a 67·6% decrease in blood insulin concentration compared to saline infusion (P<0·01); however, no glycemia changes occurred. During glucose 5·6 mM intracerebroventricular infusion, the firing rate of the vagus nerve was decreased 39% and sympathetic nerve activity was increased 177% compared to saline infusion (P<0·01). DISCUSSION: Glucose injection into the brain in the hypothalamic area modulates glucose homeostasis, which might be mediated by the sensitivity of the hypothalamic area to local changes in glucose concentration. We suggest that gluconeurons in the hypothalamus contribute to the control of glycemia through ANS activity.

Effect of prolactin acting on the coeliac ganglion via the superior ovarian nerve on ovarian function in the postpartum lactating and non-lactating rat.

Whether prolactin (PRL) has a luteotrophic or luteolytic effect in the rat ovary depends on the nature of the corpora lutea present in the ovaries and the hormonal environment to which they are exposed. The aim was to investigate the effect of PRL acting on the coeliac ganglion (CG) on the function of the corpora lutea on day 4 postpartum under either lactating or non-lactating conditions, using the CG-superior ovarian nerve-ovary system. The ovarian release of progesterone (P), estradiol, PGF2α, and nitrites was assessed in the ovarian compartment at different incubation times. Luteal mRNA expression of 3ß-HSD, 20α-HSD, aromatase, PGF2α receptor, iNOS, Bcl-2, Bax, Fas and FasL was analysed in the corpus luteum of pregnancy at the end of the experiments. Comparative analysis of control groups showed that the ovarian release of P, nitrites, and PGF2α, the expression of PGF2α receptor, and the Bcl-2/Bax ratio were lower in non-lactating rats, with increased release of estradiol, and higher expression of aromatase, Fas and FasL, demonstrating the higher luteal functionality in ovaries of lactating animals. PRL added to the CG compartment increased the ovarian release of P, estradiol, nitrites and PGF2α, and decreased the Bcl-2/Bax ratio in non-lactating rats; yet, with the exception of a reduction in the release of nitrites, such parameters were not modified in lactating animals. Together, these data suggest that the CG is able to respond to the effect of PRL and, via a neural pathway, fine-tune the physiology of the ovary under different hormonal conditions.

Protective effect of oestradiol in the coeliac ganglion against ovarian apoptotic mechanism on dioestrus.

The aims of this work were to investigate if oestradiol 10(-8)M in the incubation media of either the ovary alone (OV) or the ganglion compartment of an ex vivo coeliac ganglion-superior ovarian nerve-ovary system (a) modifies the release of ovarian progesterone (P4) and oestradiol (E2) on dioestrus II, and (b) modifies the ovarian gene expression of 3ß-HSD and 20α-HSD enzymes and markers of apoptosis. The concentration of ovarian P4 release was measured in both experimental schemes, and ovarian P4 and E2 in the ex vivo system by RIA at different times. The expression of 3ß-hydroxysteroid dehydrogenase, 20α-hydroxysteroid dehydrogenase and antiapoptotic bcl-2 and proapoptotic bax by RT-PCR were determined. E2 added in the coeliac ganglion caused an increase in the ovarian release of the P4, E2 and 3ß-HSD, while in the ovary incubation alone it decreased P4 and 3ß-HSD but increased and 20α-HSD and bax/bcl-2 ratio. It is concluded that through a direct effect on the ovary, E2 promotes luteal regression in DII rats, but the addition of E2 in the coeliac ganglion does not have the same effect. The peripheral nervous system, through the superior ovarian nerve, has a protective effect against the apoptotic mechanism on DII.

Evidence for a celiac ganglion-ovarian kisspeptin neural network in the rat: intraovarian anti-kisspeptin delays vaginal opening and alters estrous cyclicity.

Kisspeptin and its receptor GPR54 have been described as key hypothalamic components in the regulation of GnRH secretion. Kisspeptin is also present in several regions of the central nervous system and the peripheral organs and has recently been identified in the superior ganglion. Herein, we tested the possibility that ovarian kisspeptin is regulated by the sympathetic nervous system and participates locally in the regulation of ovarian function. Both ovarian and celiac ganglion kisspeptin mRNA levels increase during development, whereas kisspeptin peptide levels and plasma levels decrease during development. In the celiac ganglion, kisspeptin colocalized with tyrosine hydroxylase, indicating potential kisspeptin synthesis and transport within the sympathetic neurons. A continuous (64 h) cold stress induced marked changes within the kisspeptin neural system along the celiac ganglion-ovary axis. In vitro incubation with the ß-adrenergic agonist isoproterenol increased ovarian kisspeptin mRNA and peptide levels, and this increase was inhibited by treatment with the ß-antagonist propranolol. Sectioning the superior ovarian nerve altered the feedback information within the kisspeptin celiac ganglion-ovary axis. In vivo administration of a kisspeptin antagonist to the left ovarian bursa of 22- to 50-d-old unilaterally ovariectomized rats delayed the vaginal opening, decreased the percentage of estrous cyclicity, and decreased plasma, ovarian, and celiac ganglion kisspeptin concentrations but did not modify the LH plasma levels. These results indicate that the intraovarian kisspeptin system may be regulated by sympathetic nerve activity and that the peptide, either from a neural or ovarian origin, is required for proper coordinated ovarian function.

Estradiol promotes luteal regression through a direct effect on the ovary and an indirect effect from the celiac ganglion via the superior ovarian nerve.

There is evidence suggesting that estradiol (E(2)) regulates the physiology of the ovary and the sympathetic neurons associated with the reproductive function. The objective of this study was to investigate the effect of E(2) on the function of late pregnant rat ovaries, acting either directly on the ovarian tissue or indirectly via the superior ovarian nerve (SON) from the celiac ganglion (CG). We used in vitro ovary (OV) or ex vivo CG-SON-OV incubation systems from day 21 pregnant rats. Various concentrations of E(2 )were added to the incubation media of either the OV alone or the ganglion compartment of the CG-SON-OV system. In both experimental schemes, we measured the concentration of progesterone in the OV incubation media by radioimmunoassay at different times. Luteal messenger RNA (mRNA) expression of 3ß-hydroxysteroid dehydrogenase (3ß-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD) enzymes, respectively, involved in progesterone synthesis and catabolism, and of antiapoptotic B-cell lymphoma 2 (Bcl-2) and proapoptotic Bcl-2-associated X protein (Bax), were measured by reverse transcriptase-polymerase chain reaction (RT-PCR) at the end of the incubation period. Estradiol added directly to the OV incubation or to the CG of the CG-SON-OV system caused a decline in the concentration of progesterone accumulated in the incubation media. In addition, E(2), when added to the OV incubation, decreased the expression of 3ß-HSD and the ratio of Bcl-2/Bax. We conclude that through a direct effect on the OV, E(2) favors luteal regression at the end of pregnancy in rats, in association with neural modulation from the CG via the SON.

Involvement of the oestrogenic receptors in superior mesenteric ganglion on the ovarian steroidogenesis in rat.

Oestradiol (E(2)) is a key hormone in the regulation of reproductive processes. The aims of this work were a) to examine the distributions of oestrogen receptor α (ERα) and ERß in the neurons of the superior mesenteric ganglion (SMG) in the oestrus stage by immunohistochemistry, b) to demonstrate whether E(2) in the SMG modifies progesterone (P(4)), androstenedione (A(2)) and nitrite release in the ovarian compartment on oestrus day and c) to demonstrate whether E(2) in the ganglion modifies the activity and gene expression in the ovary of the steroidogenic enzymes 3ß-hydroxysteroid dehydrogenase (3ß-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD). The ex vivo SMG-ovarian nervous plexus-ovary system was used. E(2), tamoxifen (Txf) and E(2) plus Txf were added in the ganglion to measure ovarian P(4) release, while E(2) alone was added to measure ovarian A(2) and nitrites release. Immunohistochemistry revealed cytoplasmic ERα immunoreactivity only in the neural somas in the SMG. E(2) increased ovarian P(4) and A(2) release at 15, 30 and 60 min but decreased nitrites. The activity and gene expression of 3ß-HSD increased, while the activity and gene expression of 20α-HSD did not show changes with respect to the control. Txf in the ganglion diminished P(4) release only at 60 min. E(2) plus Txf in the ganglion reverted the effect of E(2) alone and the inhibitory effect of Txf. The results of this study demonstrate that ERα activation in the SMG has an impact on ovarian steroidogenesis in rats, thus providing evidence for the critical role of peripheral system neurons in the control of ovarian functions under normal and pathological conditions.

Androstenedione acts on the coeliac ganglion and modulates luteal function via the superior ovarian nerve in the postpartum rat.

Androstenedione can affect luteal function via a neural pathway in the late pregnant rat. Here, we investigate whether androstenedione is capable of opposing to regression of pregnancy corpus luteum that occurs after parturition, indirectly, from the coeliac ganglion. Thus, androstenedione was added into the ganglionar compartment of an ex vivo coeliac ganglion-superior ovarian nerve-ovary system isolated from non-lactating rats on day 4 postpartum. At the end of incubation, we measured the abundance of progesterone, androstenedione and oestradiol released into the ovarian compartment. Luteal mRNA expression and activity of progesterone synthesis and degradation enzymes, 3ß-hydroxysteroid-dehydrogenase (3ß-HSD) and 20α-hydroxysteroid-dehydrogenase (20α-HSD), respectively, as well as the aromatase, Bcl-2, Bax, Fas and FasL transcript levels, were also determined. Additionally, we measured the ovarian release of norepinephrine, nitric oxide and luteal inducible nitric oxide synthase (iNOS) mRNA expression. The presence of androstenedione in the ganglion compartment significantly increased the release of ovarian progesterone, androstenedione and oestradiol without modifying 3ß-HSD and 20α-HSD activities or mRNA expression. The ovarian release of oestradiol in response to the presence of androstenedione in the ganglion compartment declined with time of incubation in accord with a reduction in the aromatase mRNA expression. Androstenedione added to the ganglion compartment decreased FasL mRNA expression, without affecting luteal Bcl-2, Bax and Fas transcript levels; also increased the release of norepinephrine, decreased the release of nitric oxide and increased iNOS mRNA. In summary, on day 4 after parturition, androstenedione can mediate a luteotropic effect acting at the coeliac ganglion and transmitting to the ovary a signaling via a neural pathway in association with increased release of norepinephrine, decreased nitric oxide release, and decreased expression of FasL.

The essential oil of Croton nepetaefolius selectively blocks histamine-augmented neuronal excitability in guinea-pig celiac ganglion.

OBJECTIVES: Croton nepetaefolius is a medicinal plant useful against intestinal disorders. In this study, we elucidate the effects of its essential oil (EOCN) on sympathetic neurons, with emphasis on the interaction of EOCN- and histamine-induced effects. METHODS: The effects of EOCN and histamine were studied in guinea-pig celiac ganglion in vitro. KEY FINDINGS: Histamine significantly altered the resting potential (E(m)) and the input resistance (R(i)) of phasic neurons (from -56.6 +/- 1.78 mV and 88.6 +/- 11.43 MOmega, to -52.9 +/- 1.96 mV and 108.6 +/- 11.00 MOmega, respectively). E(m), R(i) and the histamine-induced alterations of these parameters were not affected by 200 microg/ml EOCN. The number of action potentials produced by a 1-s (two-times threshold) depolarising current and the current threshold (I(th)) for eliciting action potentials (rheobase) were evaluated. Number of action potentials and I(th) were altered by histamine (from 2.6 +/- 0.43 action potentials and 105.4 +/- 11.15 pA to 6.2 +/- 1.16 action potentials and 67.3 +/- 8.21 pA, respectively). EOCN alone did not affect number of action potentials and I(th) but it fully blocked the histamine-induced modifications of number of action potentials and I(th). All the effects produced by histamine were abolished by pyrilamine. CONCLUSIONS: EOCN selectively blocked histamine-induced modulation of active membrane properties.

Ovaric physiology in the first oestral cycle: influence of noradrenergic and cholinergic neural stimuli from coeliac ganglion.

Both peripheral innervation and nitric oxide (NO) participate in ovarian steroidogenesis. The aims of the work were (1) to investigate whether ganglionic noradrenergic (NE) and cholinergic (Ach) stimulus modify the ovarian steroids and NO release and (2) to examine the effect of those stimuli on the mRNA expression of 3beta-HSD and P450 aromatase in the ovary. The experiments were carried out using the ex vivo coeliac ganglion-superior ovarian nerve-ovary (CG-SON-O) system of rats in the first oestral cycle. The system was incubated in a buffer solution for 120min, with the ganglion and ovary located in different compartments and linked by the SON. NE and Ach were added into the ganglion compartment. Both NE and Ach predominantly induced ovarian release of androstenedione and oestradiol while inhibited progesterone release. Ovarian NO release increased after ganglionic stimulation during proestrous while its secretion decreased during the diestrous. Noteworthily, 3beta-HSD and P450 aromatase expression were modulated by neural stimulation. In the follicular phase, Ach in CG increased 3beta-HSD and NE increased P450 aromatase. In the luteal phase both neurotransmitters increased 3beta-HSD and Ach increased P450 aromatase transcript levels. All above observations suggest that the preponderancy of an either noradrenergic or cholinergic effect would depend on the stage of the first oestral cycle in the rat. The ovarian response to noradrenergic and cholinergic stimuli on GC, via SON, is strongly linked to oestral-stage-specific ovarian structures and their secretion products.

Norepinephrine modulates the effect of neuropeptides in coeliac ganglion on ovarian hormones release: its relationship with ovarian nitric oxide and nerve growth factor.

OBJECTIVE: Ovarian steroids are modulated by neural influences. In this work we investigate whether norepinephrine (NE) modifies the vasoactive intestinal peptide (VIP) or neuropeptide Y (NPY) actions in coeliac ganglion (CG) on the ovarian hormone release, and evaluate the participation of nitric oxide (NO), measured as nitrite, and of inducible nitric oxide synthetase (iNOS) protein, nerve growth factor (NGF) and its trkA receptor gene expression in the ovarian response. METHODS: The study was performed in the ex vivo CG-superior ovarian nerve (SON)-ovary system of rats on diestrus day 2 (D2). CG and ovary were placed in separate compartments connected by the SON and incubated with Krebs-Ringer buffer. After addition of 50 ng/ml VIP, 50 ng/ml NPY, 10-6 M NE, or a mix of VIP+NE or NPY+NE in ganglion, samples from the ovarian compartment were taken at different times throughout 180 minutes to measure progesterone, androstenedione and nitrite levels. RESULTS: VIP and NPY in ganglion induced an increase of progesterone release that was associated for VIP, but not NPY, with a decrease of ovarian nitrite levels, iNOS protein, and NGF/trkA receptor mRNA expression. By contrast, NE in ganglion decreased progesterone, an effect that was suppressed by addition of propranolol in ganglion, and increased nitrites/iNOS and NGF/trkA receptor expression in ovary. GABA A receptor antagonist bicuculline (20 muM) added in ovarian compartment prevented the inhibitory effect on progesterone caused by NE in CG. Androstenedione was not modified under neuropeptides or NE ganglionic stimulation. CONCLUSIONS: Finally, results from VIP+NE or NPY+NE in ganglion showed that ovarian response on D2 induced by VIP or NPY alone is moderated by the opposite action of NE, and occurs only on progesterone, the most sensitive steroid to neural action.

Allosteric interaction of the anticholinergic drug [N-(4-phenyl)-phenacyl-l-hyoscyamine] (Phenthonium) with nicotinic receptors of post-ganglionic sympathetic neurons of the rat vas deferens.

Phenthonium (Phen), a quaternary analog of hyoscyamine, is a blocker of muscarinic activity and an allosteric blocker of alpha(1)2betagammaepsilon nicotinic receptors. Specifically, Phenthonium increases the spontaneous release of acetylcholine at the motor endplate without depolarizing the muscle or inhibiting cholinesterase activity. This paper compares Phenthonium's effects on sympathetic transmission and on ganglionic nicotinic receptor activation. Neurotransmitter release and twitch of the rat vas deferens were induced either by electrical stimulation or by 1,1-dimethyl-4-phenylpiperazine (DMPP) activation of nicotinic receptors. Contractions independent of transmitter release were induced by noradrenaline and adenosine 5'-triphosphate (ATP). Phenthonium inhibited transmitter release and depressed twitch without changing the responsiveness to noradrenaline or ATP. Twitch depression did not occur after K(+)-channel blockade with 4-aminopyridine (4-AP) or charybdotoxin. DMPP had a similar effect, but high concentrations induced contraction of non-stimulated organs. Incubation of Phenthonium inhibited further DMPP twitch depression and non-competitively depressed the contractile responses elicited by DMPP. Furthermore, mecamylamine, but neither methyllycaconitine nor atropine, blocked the contraction elicited by DMPP. Phenthonium and DMPP are K(+)-channel openers that primarily inhibit sympathetic transmission. Contraction induced by DMPP was probably mediated by neuronal nicotinic receptor other than the alpha7 subtype. The blockade of DMPP contractile response was unrelated to Phenthonium's antimuscarinic or K(+)-channel opening activities. Since Phenthonium's quaternary chemical structure limits its membrane diffusion, the non-competitive inhibition of DMPP excitatory responses should be linked to allosteric interaction with neuronal nicotinic receptors that putatively qualify Phenthonium as a novel modulator of cholinergic synapses.

Androgen receptors in coeliac ganglion in late pregnant rat.

The ovarian function is controlled by endocrine factors and neural influence. In late pregnant rat, androstenedione, from the coeliac ganglion, has a luteotrophic effect in the ex vivo coeliac ganglion-superior ovarian nerve-ovary system. In this work we investigate the presence of androgen receptors in the coeliac ganglion of late pregnant rats by immunohistochemistry. We also explore, from a physiological point of view, the potential participation of these receptors in the androstenedione ganglionic action on progesterone release and metabolism, as well as on nitrites release in the ovary compartment. The coeliac ganglion was isolated after being fixed in situ and immunohistochemistry was performed. In the system, three experimental groups were used with the addition of (a) androstenedione, (b) flutamide, and (c) androstenedione plus flutamide in the ganglion compartment. Progesterone and nitrite concentrations were determined in the ovary compartment at different incubation times. Corpora lutea samples isolated at the end of incubation were used to determine the expressions and activities of the progesterone synthesis (3beta-hydroxysteroid-dehydrogenase, 3beta-HSD) and degradation (20alpha-hydroxysteroid-dehydrogenase, 20alpha-HSD) enzymes. Immunohistochemistry revealed cytoplasmatic androgen receptor immunoreactivity in neural somas in the coeliac ganglion. In the coeliac ganglion-superior ovarian nerve-ovary system, androstenedione addition increased 3beta-HSD and decreased 20alpha-HSD, showed a tendency to decrease 20alpha-HSD expression, and increased nitrites release in relation to control. Androstenedione plus flutamide decreased progesterone and nitrites release in relation to the androstenedione group. This work demonstrates the presence of androgen receptors in neurons of celiac ganglion and provides evidence for the luteotrophic action of androstenedione via a neural pathway that may be mediated by these receptors.

Prepubertal estrogen exposure modifies neurotrophin receptor expression in celiac neurons and alters ovarian innervation.

Estradiol is a key hormone in the regulation of reproductive processes acting both on peripheral organs and sympathetic neurons associated to reproductive function. However, many of its regulatory effects on the development and function on the sympathetic neurons have not been completely clarified. Sympathetic neurons located in the celiac ganglion projects to visceral, vascular and glandular targets, and contribute to ovarian innervation, being the main source of sympathetic fibers. In the present study, we analyze the effects of elevated levels of exogenous estrogen during the prepubertal period in post-ganglionic sympathetic neurons. Estrogen exposure induced a significant increase in sympathetic celiac neuronal size and modified the expression of neurotrophin receptor p75. This change affected mainly small and medium size neurons. The effect of estrogens on innervation of celiac target organs was heterogeneous, inducing a significant increase in catecholaminergic innervation of the ovary, but not of the pyloric muscular layers. These findings further support the role of estrogen as a modulator of neuronal plasticity and suggest that estrogen could modify some features involved in the relation between sympathetic immature peripheral neurons and their target organs throughout a neurotrophin-dependent mechanism.

Cardiovascular responses produced by 5-hydroxytriptamine:a pharmacological update on the receptors/mechanisms involved and therapeutic implications.

The complexity of cardiovascular responses produced by 5-hydroxytryptamine (5-HT, serotonin), including bradycardia or tachycardia, hypotension or hypertension, and vasodilatation or vasoconstriction, has been explained by the capability of this monoamine to interact with different receptors in the central nervous system (CNS), on the autonomic ganglia and postganglionic nerve endings, on vascular smooth muscle and endothelium, and on the cardiac tissue. Depending, among other factors, on the species, the vascular bed under study, and the experimental conditions, these responses are mainly mediated by 5-HT(1), 5-HT(2), 5-HT(3), 5-HT(4), 5-ht(5A/5B), and 5-HT(7) receptors as well as by a tyramine-like action or unidentified mechanisms. It is noteworthy that 5-HT(6) receptors do not seem to be involved in the cardiovascular responses to 5-HT. Regarding heart rate, intravenous (i.v.) administration of 5-HT usually lowers this variable by eliciting a von Bezold-Jarisch-like reflex via 5-HT(3) receptors located on sensory vagal nerve endings in the heart. Other bradycardic mechanisms include cardiac sympatho-inhibition by prejunctional 5-HT(1B/1D) receptors and, in the case of the rat, an additional 5-ht(5A/5B) receptor component. Moreover, i.v. 5-HT can increase heart rate in different species (after vagotomy) by a variety of mechanisms/receptors including activation of: (1) myocardial 5-HT(2A) (rat), 5-HT(3) (dog), 5-HT(4) (pig, human), and 5-HT(7) (cat) receptors; (2) adrenomedullary 5-HT(2) (dog) and prejunctional sympatho-excitatory 5-HT(3) (rabbit) receptors associated with a release of catecholamines; (3) a tyramine-like action mechanism (guinea pig); and (4) unidentified mechanisms (certain lamellibranch and gastropod species). Furthermore, central administration of 5-HT can cause, in general, bradycardia and/or tachycardia mediated by activation of, respectively, 5-HT(1A) and 5-HT(2) receptors. On the other hand, the blood pressure response to i.v. administration of 5-HT is usually triphasic and consists of an initial short-lasting vasodepressor response due to a reflex bradycardia (mediated by 5-HT(3) receptors located on vagal afferents, via the von Bezold-Jarisch-like reflex), a middle vasopressor phase, and a late, longer-lasting, vasodepressor response. The vasopressor response is a consequence of vasoconstriction mainly mediated by 5-HT(2A) receptors; however, vasoconstriction in the canine saphenous vein and external carotid bed as well as in the porcine cephalic arteries and arteriovenous anastomoses is due to activation of 5-HT(1B) receptors. The late vasodepressor response may involve three different mechanisms: (1) direct vasorelaxation by activation of 5-HT(7) receptors located on vascular smooth muscle; (2) inhibition of the vasopressor sympathetic outflow by sympatho-inhibitory 5-HT(1A/1B/1D) receptors; and (3) release of endothelium-derived relaxing factor (nitric oxide) by 5-HT(2B) and/or 5-HT(1B/1D) receptors. Furthermore, central administration of 5-HT can cause both hypotension (mainly mediated by 5-HT(1A) receptors) and hypertension (mainly mediated by 5-HT(2) receptors). The increasing availability of new compounds with high affinity and selectivity for the different 5-HT receptor subtypes makes it possible to develop drugs with potential therapeutic usefulness in the treatment of some cardiovascular illnesses including hypertension, migraine, some peripheral vascular diseases, and heart failure.