Chronic welfare restrictions and adrenal gland morphology in broiler chickens.

Gait problems constitute an important and chronic welfare restriction for broiler chickens. The objective of the present study was to determine if adrenal gland morphology indicates chronic welfare restrictions in broiler chickens, using gait problems as the stressor. Sixty-six birds raised on a commercial unit were selected at 40 d of age and separated into groups according to gait score. One group was apparently healthy birds (AH) with gait scores of 0 to 2, and the other group had birds with gait problems (GP) that showed gait scores of 4 to 5. Birds were slaughtered and weighed, and their adrenal glands were measured and weighed; proportions of medullary and adrenocortical tissues, and lymphatic tissue and blood vessels were studied. GP birds had lower BW when compared to AH birds, and when adrenal gland weight values were adjusted to BW, a greater relative adrenal weight was observed for the GP group. Adrenals from GP birds also presented a higher proportion of blood vessels when compared to AH birds. These results might indicate increased adrenal activity and evidence of the inflammatory process as a consequence of chronic stress. Results showed that gait problems caused significant adrenal gland changes, suggesting a possible role for the study of adrenal gland morphology as an indicator of chronic welfare problems in broiler chickens.

Programming of rat adrenal medulla by neonatal hyperleptinemia: adrenal morphology, catecholamine secretion, and leptin signaling pathway.

Leptin serum concentration in early life is an important factor for adequate future development of the offspring. Previously, we demonstrated that hyperleptinemia on lactation programmed for hyperleptinemia, central leptin resistance with lower expression of the long form of leptin receptor at hypothalamus, and higher medullary catecholamine levels with cardiovascular consequences at adulthood. The central objective of this study was to determine the direct effect of leptin on adrenal medullary function of adult rats that were leptin treated during lactation. Adrenal morphology was also accessed. Recombinant murine leptin was injected in the pups during the first 10 days of life (group L, leptin-programmed) or at adulthood during 6 days (group LC). The controls of both experiments received saline (groups C and CC). Both treatments resulted in hyperleptinemia at 150 days old (+78% and 2-fold increase, respectively; P < 0.05). Programmed animals showed hypertrophy of adrenal and higher adrenal catecholamine content at 150 days old (3-fold increase, P < 0.05), and no changes were observed in the LC group. However, LC rats had lower adrenal content of tyrosine hydroxylase (-17%, P < 0.05). Leptin-programmed rats had a lower response to leptin in vitro stimulation (-22%, P < 0.05) and lower expression of key proteins of the leptin signaling pathway, leptin receptor and janus tyrosine kinase 2 in the medullas (-61% and -29%, respectively, P < 0.05). However, they presented higher expression of phosphorylated signal transducer and activator of transcription 3 (+2-fold, P < 0.05). Leptin treatment at adulthood did not affect these parameters. The higher catecholamine synthesis and secretion in the leptin-programmed rats observed in our previous study does not seem to be a consequence of the direct effect of leptin on the medullas. We suggest that the hyperleptinemia of the programmed animals increases adrenal medullary function through sympathetic nervous system activation. In conclusion, high leptin levels on lactation program the activity of the sympathoadrenal system at adulthood that may contribute to the development of adult chronic diseases such as hypertension.

Molecular imaging of adrenal gland by desorption electrospray ionization mass spectrometry.

Subtle differences in the spatial distributions of closely related compounds including norepinephrine and epinephrine as well multiple lipids are easily distinguished in adult porcine (17 x 8 mm) and rabbit (7 x 4 mm) adrenal glands in a DESI-MS imaging experiment at atmospheric pressure with a spatial resolution of approximately 200 microm. Sensitive and specific detection in the course of DESI imaging discloses details of catecholamine distribution in porcine adrenal medulla and cortex; the average mass of epinephrine interrogated in each pixel is estimated to be about 150 pg. The distribution of ascorbic acid was revealed in the negative ion mode. In addition, the distribution of cholesterol, which cannot be observed using conventional DESI, was obtained using in situ reaction with betaine aldehyde added to the DESI spray while imaging the porcine adrenal gland tissue. Four characteristic types of distributions were observed, with major amounts of the components in the medulla, the cortex, the reticular zone or in the fourth case, being homogeneously distributed. The results agree with and extend information available from histological studies.

Immunohistochemical localization of catecholamine biosynthetic enzymes in the adrenal gland of the domestic fowl (Gallus domesticus).

The present study investigated the cellular localization of 3 catecholamine biosynthetic enzymes, tyrosine hydroxylase (TH), dopamine beta-hydroxylase (DBH), and phenylethanolamine N-methyltransferase (PNMT) to identify and analyze the localization of norepinephrine (NE) and epinephrine (E) cells in the adrenal gland in the chicken using peroxidase-antiperoxidase immunohistochemical techniques. Tyrosine hydroxylase immunoreactivity (IR) was observed in almost all adrenal medullary cells of the adult chicken. Dopamine beta-hydroxylase IR coincided with that of TH. Many medullary cells also exhibited PNMT IR, but PNMT-immunonegative cells were also observed. Tyrosine hydroxylase IR was localized in the E- and NE-containing cells, but PNMT IR was localized only in the E-containing cells. Approximately 69% of medullary cells were E-containing, and the remaining were NE-containing cells. The ratio of E- and NE-containing cells between the subcapsular and central zone was statistically significant (P < 0.01). Although cortical cells of the adrenal gland did not show TH-, DBH-, or PNMT-positive reactions, ganglia close to the adrenal gland showed TH, DBH, and PNMT immunoreactivities. These findings indicated the cellular localization of 3 catecholamine-biosynthesizing enzymes in chicken adrenal medulla and suggest that the majority of medullary cell are E-containing cells. The ratio of E cells to NE cells varies among the 3 zones in the adrenal glands of the chicken.

Morphology and ultrastructure of the adrenal gland in Bactrian camels (Camelus bactrianus).

In the present study, we examined the morphological features of the adrenal gland in Bactrian camel by means of digital anatomy, light and electron microscopy. Our findings testified that the gland was divided into three parts, capsule, cortex and medulla from outside to inside as other mammals, and the cortex itself was further distinguished into four zones: zona glomerulosa, zona intermedia, zona fasciculate and zona reticularis. Notably, the zona intermedia could be seen clearly in the glands from females and castrated males, whereas it was not morphologically clear in male. There was a great deal of lipid droplets in the zona fasciculate, while it was fewer in the zona glomerulosa and zona reticularis. The cytoplasm of adrenocortical cell contained rich mitochondria and endoplasmic reticulum. The adrenal medulla was well-developed with two separations of external and internal zones. The most obvious histological property of adrenal medulla cells were that they contained a huge number of electron-dense granules enveloped by the membrane, and so medulla cells could be divided into norepinephrine cells and epinephrine cells. Moreover, the cortical cuffs were frequently present in adrenal gland. Results of this study provides a theoretical basis necessary for ongoing investigations on Bactrian camels and their good adaptability in arid and semi-arid circumstances.

Up-regulation of adrenal cortical and medullary atrial natriuretic peptide and gene expression in rats with deoxycorticosterone acetate-salt treatment.

Our previous study demonstrated that human adrenal medulla is a site of atrial natriuretic peptide (ANP) synthesis. To further evaluate the role of adrenal ANP in body fluid homeostasis, we investigated the changes in adrenal ANP in rats receiving deoxycorticosterone acetate (DOCA)-salt treatment. In situ hybridization and immunohistochemical study showed that adrenal ANP messenger RNA (mRNA) and ANP-like immunoreactivities (ANP-LI) were mainly localized in the zona glomerulosa and medulla of vehicle-treated rats. DOCA-salt treatment activated ANP mRNA and peptide expression in all adrenal zones, especially in the zona fasciculata/reticularis from 12 h to the entire 8-day study period. Using a semiquantitative RT-PCR technique, the relative quantities of ANP mRNA in the adrenals of the DOCA-salt-treated group were significantly increased from 1 to 8 days, whereas the adrenal weights of DOCA-salt-treated rats were significantly decreased from day 2 to day 8. Our results are the first to indicate that ANP is synthesized not only in the adrenal medulla but also in the adrenal cortex and their syntheses are markedly increased in DOCA-salt-treated rats. These results imply that adrenal ANP may participate in the intraadrenal regulation of adrenal function on water-electrolyte homeostasis in an autocrine or paracrine manner.

Reduced pain-related behavior by adrenal medullary transplants in rats with experimental painful peripheral neuropathy.

Adrenal medullary transplants in the spinal subarachnoid space, by providing a continual source of opioid peptides and catecholamines, offer a potentially important adjunct in the management of chronic pain. While previous studies have shown that this approach is effective against high-intensity phasic stimuli, adrenal medullary implants need to be evaluated against long-term and abnormal pain syndromes before transplantation can be used for human chronic pain. Using a recently developed model of painful peripheral neuropathy, the effects of adrenal medullary chromaffin cells transplanted into the subarachnoid space was evaluated. Peripheral mononeuropathy was induced by loosely tying 4 ligatures (4-0 chromic gut) around the right sciatic nerve. This procedure produces various pain symptoms including allodynia, hyperalgesia and dysesthesia. Rats were given either adrenal medullary tissue or control striated muscle transplants. Animals with adrenal medullary tissue transplants showed markedly decreased allodynia to innocuous cold as early as 1 week post-transplantation. In addition, hyperalgesia to a noxious thermal stimulus was eliminated by adrenal medullary, but not control, transplants. Touch-evoked allodynia was only slightly reduced by adrenal medullary transplants. In addition, indicators of spontaneous pain appeared reduced in animals with adrenal medullary transplants. These findings indicate that adrenal medullary transplants may be effective in reducing neuropathic pain.

Increased expression of tenascin in pheochromocytomas correlates with malignancy.

Tenascin is a significant extracellular matrix glycoprotein, which is upregulated in various neoplasias and pathologic processes. Pheochromocytomas are rare tumors of the sympathoadrenal system, whose malignancy is almost impossible to predict. There are no histologic or chemical markers available that would define the malignant behavior of these tumors, except the discovery of metastases. In our search for new markers, we investigated the immunohistochemical expression of tenascin in a large number of pheochromocytomas and paragangliomas. Seven tumors were metastasized and were thus considered malignant. Normal adrenal medulla was tenascin negative. A striking difference was seen between malignant and benign pheochromocytomas. All malignant pheochromocytomas expressed stromal tenascin strongly or moderately, whereas most benign pheochromocytomas (28 of 37, 70%) showed no or only weak immunopositivity. The staining was strong or moderate also in 13 of 28 (46%) of the tumors that showed histologically suspicious features, here called borderline tumors. Paragangliomas showed a more heterogeneous staining pattern, and no significant difference was found between benign and malignant paragangliomas. To our knowledge, this is the first study to demonstrate the expression of tenascin in pheochromocytomas and particularly the enhanced expression in malignant pheochromocytomas. We therefore suggest that tenascin may be associated with the malignant transformation and metastasis of pheochromocytomas. It is also a potential marker predicting more aggressive behavior in pheochromocytomas.

Absence of appreciable tolerance and morphine cross-tolerance in rats with adrenal medullary transplants in the spinal cord.

Adrenal medullary transplants in the spinal subarachnoid space may be a means of achieving sustained local delivery of pain-reducing neuroactive substances on a continually renewable basis. However, a potential limitation of this approach is tolerance development to agents released from the transplanted cells. In particular, since adrenal medullary chromaffin cells release opioid peptides, reduced antinociceptive efficacy of opioids is possible. To determine this, alterations in the dose-effectiveness of morphine were assessed in animals with adrenal medullary transplants. Results indicated that, not only was there no apparent tolerance, but that adrenal medullary transplants could potentiate the analgesic efficacy of morphine. An additional goal of these studies was to determine whether chronic or intermittent nicotine could produce increased antinociception, since stimulation of cell surface nicotinic receptors increases release of neuroactive substances from chromaffin cells. This was assessed using subcutaneously implanted nicotine pellets or repeated systemic administration of nicotine. Findings indicated that exposure to nicotine results in an acute tolerance, or tachyphylaxis, to nicotine which is rapidly reversed following cessation of nicotinic stimulation. Together, these results suggest that adrenal medullary transplants may provide a constant source of opioid peptides, augmentable by intermittent nicotinic stimulation, without the development of appreciable tolerance to these pain-reducing neuroactive substances.

Adrenal medullary ganglion neurons project into the splanchnic nerve.

Retrograde tract-tracing was used to study the projections of adrenal medullary ganglion neurons. The splanchnic nerve was cut close to the suprarenal ganglia and the retrograde tracer FluoroGold was applied at the site of nerve transection. Groups of adrenal medullary ganglion neurons exhibited FlurorGold- or Fast Blue-induced fluorescence restricted to the perikarya. Using immunohistochemistry most retrogradely labelled ganglion neurons showed immunoreactivity for neuropeptide Y. In addition, after splanchnicotomy most ganglion neurons expressed galanin and galanin message-associated peptide immunoreactivities which could not be observed in control adrenals. Taken together, the present results strongly indicate that adrenal medullary ganglion neurons project back into the splanchnic nerve perhaps representing feedback system modulating the preganglionic innervation of the adrenal gland.

A quantitative analysis of rat adrenal chromaffin tissue: morphometric analysis at tissue and cellular level correlated with catecholamine content.

A morphometric analysis of normal Wistar rat adrenal medulla following perfusion fixation and Araldite embedding, was correlated with catecholamine levels on fresh tissue, measured by high-performance liquid chromatography. The mean volume of whole adrenal is 13.2 mm3 and the mean medullary volume 1.3 mm3. Volume density estimates showed that the medulla is composed of 63% chromaffin tissue with an adrenaline to noradrenaline storing cell ratio of 4.4:1. The vasculature occupies 20%, neuronal tissue 5% and interstitial tissues 12% of the medulla. A comparison was made of cell volumes, cell numbers and volume and surface density estimates of cytoplasmic organelles in adrenaline and noradrenaline storing cells. The mean cell volume of adrenaline storing cells at 1300 micron3 is larger than that of noradrenaline storing cells at 980 micron3. A single adrenal medulla contains 4.4-5.7 X 10(5) adrenaline cells and 1.5-1.9 X 10(5) noradrenaline cells. Chromaffin granules account for approximately 30% of the volume of the cytoplasm; the numerical density of granules at different sites in the cell was calculated for adrenaline cells. The volume density of mitochondria (4%) and the surface density of mitochondrial membranes (the ratio of outer to inner membrane being approximately 1:2.3) were similar in both cell types. Rough endoplasmic reticulum was the only organelle to show a significant difference in volume and surface density between the two cell types. Adrenaline storing cells have stacks of rough endoplasmic reticulum which have two to three times the surface and volume densities of that found diffusely scattered throughout noradrenaline cells. The adrenaline content of an adrenaline storing cell is 0.14 X 10(-6) microM and that of a granule 3.0 X 10(-12) or 3.8 X 10(-12) mumoles depending on the method of calculation. The noradrenaline content of noradrenaline storing cells can only be calculated on the assumption that all noradrenaline is stored in this cell type though it is likely that some is contained within adrenaline cells. Based on this assumption the noradrenaline content is 0.17 X 10(-6) mumoles per cell and 5 X 10(-12) mumoles per granule. The present study provides baseline morphometric data on the rat adrenal medulla at tissue and cellular level correlated with amine levels in adrenaline and noradrenaline storing cells and granules.

Action of 6-hydroxydopamine on lamb sympathetic ganglia, vas deferens and adrenal medulla: a combined histochemical, ultrastructural and biochemical comparison with the effects of reserpine.

1. The effects of a single dose of 6-hydroxydopamine (6-OHDA) compared with those of chronic reserpine treatment were studied in lamb sympathetic neurones and adrenal medulla by a combination of fluorescence histochemistry, electron microscopy and radiochemical assay.2. In sympathetic ganglia, 6-OHDA produced a rise in noradrenaline concentration within 24 h, and falls in tyrosine hydroxylase and monoamine oxidase activities, whereas reserpine caused a fall in noradrenaline, a rise in tyrosine hydroxylase activity and no change in monoamine oxidase activity. The fluorescence of intra- and postganglionic axons increased greatly within 24 h of 6-OHDA, and there was a corresponding accumulation of large dense-core vesicles within many axons whose neurotubules were disrupted. The changes were almost reversed after 3 weeks.3. In the vas deferens, the concentration of noradrenaline and tyrosine hydroxylase and monoamine oxidase activities had all fallen 24 h after 6-OHDA treatment and had started to recover 3 weeks later. In the adrenal medulla, 6-OHDA did not alter NA concentrations but increased tyrosine hydroxylase activity whereas reserpine depleted noradrenaline and increased tyrosine hydroxylase activity.4. The changes produced in sympathetic ganglia by 6-OHDA may be due both to a direct action on the axoplasmic transport of noradrenaline containing vesicles and indirectly to the reaction of the neurones to loss of the integrity of their axons.

Stereological estimation of number and volume of pulmonary neuroepithelial bodies (NEBs) in neonatal hamster lungs.

The number and volume of pulmonary neuroepithelial bodies (NEBs) of 1- and 4-week-old hamsters were estimated using unbiased stereological principles and systematic sampling techniques. For comparative purposes, volume estimations were also made in the carotid body, the parathyroid gland, and the adrenal medulla. A significant decrease was found in the total number of NEBs, immunoreactive for CGRP, between 1 and 4 weeks. Individual as well as cumulative NEB volume also decreased significantly. The cumulative NEB volume in 1-week-old hamsters was in the same range as the volumes of the carotids and parathyroids in the same animals. The postnatal decrease of the NEB number suggests that the NEBs are of primary potential importance in the neonatal stage, when they may complement the chemoreceptor function of the carotid bodies, which are relatively inactive at birth. Since the cumulative NEB volume (at least at the age of 1 week) is equal to that of the carotid bodies and the parathyroids, their physiological function may be of similar importance.

Effect of exercise on mRNA expression of select adrenal medullary catecholamine biosynthetic enzymes.

The effect of submaximal endurance training (SET) on sympathoadrenal activity is not clear. We tested the hypothesis that SET (90 min/day, 5 days/wk, for 12 wk) elevates mRNA expression of catecholamine (CA) biosynthetic enzymes, tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DbetaH) in the adrenal medullae of adult, female Sprague-Dawley rats. SET increased TH protein level by 35%, TH activity by 62%, TH mRNA expression by 40%, and DbetaH mRNA expression by 67%. In addition, we examined the effect of SET on Fos-related antigens (FRAs), FRA-2 immunoreactivity, and activator protein (AP)-1 binding activity. SET increased AP-1 binding activity by 78%; however, it did not affect late FRAs and FRA-2 immunoreactivity. Because the regulation of neuropeptide Y (NPY) often parallels that of CAs, we also examined the effect of SET on NPY mRNA expression. Indeed, SET elevated NPY mRNA expression as well. We conclude that 1) SET elicits a pretranslational stimulatory effect on adrenomedullary CA biosynthetic enzymes, 2) another immediate early mRNA product, rather than FRA-2, may contribute to the increase in AP-1 binding activity in response to SET, and 3) SET increases NPY mRNA expression.

Quantitative autoradiography of angiotensin II AT2 receptors with [125I]CGP 42112.

Most radiolabeled ligands for angiotensin II (Ang II) receptors do not discriminate between the AT1 and AT2 receptor subtypes, which must be distinguished by displacement with selective AT1 or AT2 ligands. We compared [125I]CGP 42112 with the non-selective agonist [125I]Sar1 Angiotensin II. We studied the inferior olive, medial geniculate nucleus and the adrenal medulla, areas rich in AT2 receptors, using both ligands with quantitative autoradiography and membrane binding techniques. [125I]CGP 42112 bound with high affinity (Kd = 0.07-0.3 nM, depending on the area studied). [125I]CGP 42112 binding was selective for AT2 receptors, as determined by lack of competition with the AT1 ligand losartan, and competition by the AT2 ligands PD 123177 and unlabeled CGP 42112 and the non-selective peptides Ang II and angiotensin III (Ang III). Using [125I]CGP 42112 binding, we found the same order of potency: CGP 42112 > Ang II = Ang III > PD 123177 using both quantitative autoradiography or membrane binding methods. Our results demonstrate that [125I]CGP 42112 is the most selective, highest affinity ligand available for AT2 receptors. Because of these characteristics, and low non-specific binding, quantitative autoradiography with [125I]CGP 42112 is the method of choice to selectively characterize AT2 receptors, especially in tissues like the brain, with a highly heterogeneous distribution of receptor subtypes.

The non-neurogenic catecholamine response of the fetal adrenal to hypoxia is dependent on activation of voltage sensitive Ca2+ channels.

We have investigated the cellular mechanisms underlying the catecholamine response of the fetal sheep adrenal to hypoxia before and after the development of adrenal innervation. Adrenals were collected before (80-100 days gestation: n = 7) and after (135-146 days gestation: n = 10) development of innervation and retrogradely perfused with oxygenated Krebs bicarbonate buffer in vitro via the renal vein. Adrenal hypoxia was induced by perfusion with hypoxic Krebs buffer (pO2 = 46.7 +/- 2.4 mm Hg) for 30 min periods in the presence and absence of hexamethonium (500 microM), Ca2+ (2.5 mM), nifedipine (1 microM) and KCl (10 mM). Hypoxia stimulated an increase (P < 0.001) in the output of noradrenaline at 80-100 days (3 min pre hypoxia, 0.18 +/- 0.07 nmol/3 min; 20 min hypoxia, 0.74 +/- 0.22 nmol/3 min) and at 135-146 days (3 min pre hypoxia, 0.53 +/- 0.20 nmol/3 min; 20 min hypoxia, 1.71 +/- 0.85 nmol/3 min). Adrenaline output was also higher (P < 0.001) than basal values (80-100 days, 0.11 +/- 0.06 nmol/3 min; 135-146 days, 0.53 +/- 0.15 nmol/3 min) after 20 min hypoxia (0.41 +/- 0.20 nmol/3 min and 1.35 +/- 0.56 nmol/3 min respectively). The catecholamine responses to hypoxia were abolished by removal of Ca2+ from the adrenal perfusate. There was a reduction (P < 0.05) in the catecholamine secretory response to hypoxia in the presence of nifedipine. Noradrenaline output decreased from 4.33 +/- 0.84 nmol/30 min to 0.16 +/- 0.49 nmol/30 min and adrenaline output decreased from 3.16 +/- 1.66 nmol/30 min to -0.01 +/- 0.24 nmol/30 min in the presence of nifedipine. The fetal adrenal secretes catecholamines by a direct or non-neurogenic mechanism in response to hypoxia. This secretory response is dependent on the activation of voltage sensitive Ca2+ channels in the chromaffin cell membrane.

Progress in microfluorometric identification of monoamine fluorophores.

Techniques for the microfluorometric identification of monoamine fluorophores have been put forward along two different lines: (1) extension of the excitation range into the far UV and (2) computer-assisted on-line scanning recordings in combination with an automatic microfluorometric identification of fluorophores. By an extension of the excitation range to 250 nm it is possible to distinguish fluorophores of DOPA from those of dopamine, fluorophores of noradrenaline from those of adrenaline, and fluorophores of 5-OH-tryptophan from those of 5-OH-tryptamine. In a computer-assisted on-line scanning procedure using a high-aperture inverted illuminating microscope equipped with UV transmitting optics the specific formaldehyde-induced catecholamine fluorescence is characterized microfluorometrically by corrected ratios of excitation maxima (370:320 nm and 320:275 nm). Excitation data are plotted out with corresponding coordinates in the x- and in the y-axis. Furthermore the digitized values of fluorescence intensities obtained with the scanning procedure are processed according to principles of automatic image-analysis procedures.

Adrenal demedullation and peripheral 6-OHDA administration in the rabbit: effects on body weight, general activity and cardiovascular responsivity.

Two experiments were undertaken to determine the effects of systemic 6-hydroxydopamine (6-OHDA) administration and adrenal enucleation on body weight, brain and heart norepinephrine, general activity, and cardiovascular responsivity in the rabbit. 6-Hydroxydopamine produced long lasting, dose related decreases in baseline blood pressure (BP) and in pressor responses following electric shock. Partial reversal of the BP changes was observed in nonenucleated animals tested 20 days after 6-OHDA administration but not in enucleated animals tested 5, 10 or 20 days post-injection or in control animals tested 5, 10 or 15 days after vehicle injection. Although 6-OHDA administration produced transient weight loss and activity decrements, no long term weight or activity changes were observed.

Characterization of the rat adrenal medulla cultured in vitro.

A wide variety of experimental animal models have been used to investigate the mechanisms of synthesis, storage, and release of catecholamines. Whereas in vivo experimental models are situated at one end of the spectrum, cell culture models are situated at the other end. In the present study, we have characterized various aspects of the rat adrenal medulla cultured in vitro as a whole tissue, aiming to establish a new experimental model in between in vivo animal models and cell culture models. We adapted a bottle rotator system commonly used for culturing rodent whole embryos. Changes in histology, activities and mRNA levels of catecholamine-synthesizing enzymes, and concentrations of catecholamines in the adrenal medulla were studied. In addition, the effects of cholinergic stimulation on catecholamine release from the adrenal medulla were examined. Overall the results indicate that various aspects of the adrenal medulla become stable after 4 d of culture and the adrenal medulla at this stage releases catecholamines in response to cholinergic stimulation. The whole adrenal medulla culture system may be a useful tool for investigating catecholamine-related functions dependent on intercellular reactions or communications.

Altered gonadal hormone level and constant light-induced stress interfere with the response of the adrenal medulla to oxytocin.

The effect of oxytocin (0.25 IU/100 g per day) on the adrenal medulla was examined in intact, intact estrogen-treated, castrated and castrated testosterone-treated adult male Wistar rats. Stereological analysis of the gland (N = 5 rats per group) revealed that in intact animals the number of chromaffin cells (x10(3)) was significantly increased after 3-day (saline: 467.6 +/- 27.4; oxytocin: 567.6 +/- 28.9) or 7-day (saline: 486.2 +/- 39.1; oxytocin: 618.7 +/- 36.8) oxytocin administration. During 7 days of recovery after the 7-day treatment, the chromaffin cell number returned to the control level (saline: 491.4 +/- 12.6; oxytocin: 554.4 +/- 28.7). The effect of oxytocin on chromaffin cell number was also observed in rats simultaneously injected with estradiol (0.3 micrograms/100 g per day) for 10 days (estradiol: 454.3 +/- 32.8; estradiol+oxytocin: 576.1 +/- 25.0), as well as in 10-day castrated rats (saline: 594.7 +/- 22.7; oxytocin: 765.3 +/- 33.1). Testosterone replacement (0.6 mg/100 g per day) abolished the medullary response to oxytocin (testosterone+saline: 528.5 +/- 24.7; testosterone+oxytocin: 620.8 +/- 56.0). There was a 20% rise in adrenal dopamine content (from 0.236 +/- 0.015 to 0.283 +/- 0.015 microgram per pair of glands; N = 9-12) in intact rats injected with oxytocin for 3 days. Oxytocin had no effect on any of the catecholamine levels in adrenal glands of rats exposed to stress induced by constant lighting. The present data indicate that the proliferative response of chromaffin tissue to oxytocin depends on the gonadal hormone level and the basal activity of the adrenal medulla.