Quantitative changes of CRF-like immunoreactivity in eels treated with reserpine and cortisol

Immunocytochemical techniques were applied to brain and pituitary sections of European eels after experimental manipulation of the pituitary-interrenal activity. A corticotropin-releasing factor (CRF) antiserum allowed the identification of a CRF-like peptide in the preoptic nucleus (PON) and rostral and caudal neurohypophysis (NH). CRF-immunoreactivity (ir) was not affected in solvent-injected eels compared to noninjected eels. Reserpine induced a stimulation of the pituitary interrenal axis, decreased ir-CRF in the rostral NH, but did not affect hypothalamic ir-CRF. Cortisol reduced the immunostaining of hypothalamic CRF-ir perikarya and perikarya cross-sectional area. In the rostral NH, CRF-ir fibers decreased in number and almost disappeared in long-term treated eels. The immunostaining of ACTH cells with ACTH antiserum was greatly reduced. These data suggest that cortisol induces a marked reduction in the activity of the CRF-corticotrop axis. The intensity of the ir-CRF staining observed in the caudal NH, close to the intermediate lobe (IL) was not significantly affected in reserpine-treated eels, and only slightly reduced in long-term cortisol-treated eels. The intensity of ir-CRF staining in the caudal NH did not correlate with melanocorticotropic activity or plasma cortisol level. These data suggest that immunoreactive CRF fibers in the rostral and caudal NH are differently regulated.     

Immunocytochemical identification of CRF-like and SRIF-like peptides in the brain and the pituitary of cyprinid fish

Summary Single and double immunocytochemical techniques were applied to the brain and pituitary of carps and goldfish. With the use of antiserum raised against synthetic corticotropin-releasing factor (CRF), immunoreactive perikarya were observed in the nucleus praeopticus and the nucleus praeopticus periventricularis. CRF-like-immunoreactive hypothalamic nerve fibers reach the pituitary. In cyprinids, some fine fibers enter the rostral neurohypophysis bordered by prolactin- and ACTH cells. Other thicker fibers extend ventrocaudally into the neurointermediate lobe. This CRF-like system appears to differ from the SRIF-like system, which is restricted to the proximal pars distalis of the pituitary containing somatotrophs.The technical assistance of Jacqueline Olivereau, Biologiste-Adjointe from the CNRS, is gratefully acknowledged     

Immunocytochemical localization of somatostatin and vasotocin in the brain of the Pacific hagfish,Eptatretus stouti

Summary The brain of the Pacific hagfish, Eptatretus stouti, was studied immunocytochemically using antisera against somatostatin (SRIH), arginine vasopressin (AVP), and adrenocorticotropic hormone (ACTH). SRIH-immunoreactive perikarya were distributed bilaterally in the postoptic nucleus and in the hypothalamic nucleus. Although several short, stained fibers were observed in the vicinity of the perikarya, SRIH-immunoreactivity was not found in the neurohypophysis, nor in other parts of the brain. On the other hand, presumed arginine vasotocin (AVT) perikarya were distributed in an arc-shaped region extending from the posterior part of the preoptic nucleus to the anterior-most end of the hypothalamic nucleus and projected their fibers to the neurohypophysis. Most presumptive AVT perikarya were located close to the paired prehypophysial arteries near the anterior end of the postoptic nucleus. In the neurohypophysis, abundant presumptive AVT-fibers terminated in the posterior dorsal wall, although some fibers terminated in the anterior dorsal wall and only a few fiber endings were found in the ventral wall. No ACTH-positive cells were detected in the hagfish brain or in the pituitary gland.Supported from a grant from the National Science Foundation PCM 8141393     

The distribution of corticotropin-releasing factor-immunoreactive neurons and nerve fibers in the brain of the snake,Natrix maura

Summary The anatomical distribution of neurons and nerve fibers containing corticotropin-releasing factor (CRF) has been studied in the brain of the snake, Natrix maura, by means of immunocytochemistry using an antiserum against rat CRF. To test the possible coexistence of CRF with the neurohypophysial peptides arginine vasotocin (AVT) and mesotocin (MST) adjacent sections were stained with antisera against the two latter peptides. CRF-immunoreactive (CRF-IR) neurons exist in the paraventricular nucleus (PVN). In some neurons of the PVN, coexistence of CRF with MST or of CRF with AVT has been shown. Numerous CRF-IR fibers run along the hypothalamo-hypophysial tract and end in the outer layer of the median eminence. In addition, some fibers reach the neural lobe of the hypophysis. CRF-IR perikarya have also been identified in the following locations: dorsal cortex, nucleus accumbens, amygdala, subfornical organ, lamina terminalis, nucleus of the paraventricular organ, nucleus of the oculomotor nerve, nucleus of the trigeminal nerve, and reticular formation. In addition to all these locations CRF-IR fibers were also observed in the lateral septum, supraoptic nucleus, habenula, lateral forebrain bundle, paraventricular organ, hypothalamic ventromedial nucleus, raphe and interpeduncular nuclei.     

Localization of corticotropin-releasing factor immunoreactivity in the brain of the teleost Sparus aurata

The distribution of perikarya and fibers containing corticotropin-releasing factor (CRF) was studied in the brain of the teleost Sparus aurata by immunocytochemistry using the peroxidase-antiperoxidase method. Antisera against rat CRF, arginine vasotocin, and human adrenocorticotropin (ACTH) were used. Most CRF-immunoreactive neurons were located in the nucleus lateralis tuberis, but they were absent from the nucleus preopticus, which only contained arginine vasotocin neurons. Few CRF perikarya were identified in the nucleus preopticus periventricularis and in the mesencephalic tegmentum. A conspicuous bundle of immunoreactive fibers ran along the diencephalic floor and pituitary stalk to end near the cells of the hypophysial pars intermedia. No CRF was seen near the adenohypophysial rostral pars distalis. Our results suggest that, in Sparus aurata, CRF is a releasing factor for melanotropic cells. Its role as a releasing factor for ACTH is discussed.     

Immunocytochemical distribution of corticotropin-releasing factor in the brain and hypophysis of larval Bufo arenarum; effect of KClO4 during early development

The maturation of the corticotropin-releasing factor (CRF) neuronal system was evaluated by immunocytochemistry and morphometry in Bufo arenarum, during spontaneous metamorphosis and in tadpoles with inhibited thryroid function. The first appearance of CRF immunoreactive fibers was at the end of premetamorphosis (stage VIII). These fibers were found in small numbers and weakly stained in the median eminence and infundibular stalk. With the advance of larval development, CRF-like material stained intensely and tended to aggregate in the external zone of the median eminence. At climax stages, immunoreactive fibers and perikarya (weakly stained) were identified in the interpeduncular nucleus and in the dorsal infundibular nucleus. Morphometric and immunocytochemical results indicate that the maturation of the CRF neuronal system in Bufo arenarum occurs just before metamorphic climax, coinciding with high levels of thyroid and steroid hormones. We have also found that in larvae with inhibited thyroid function, the CRF neuronal system is able to develop, and that thyroid hormone could exert a negative feedback control on the synthesis of CRF.     

Localization of corticotropin-releasing hormone (CRF)-like immunoreactivity in the central nervous system of the elasmobranch fish,Scyliorhinus canicula

Summary The occurrence and localization of immunoreactive corticotropin-releasing factor (CRF) in the brain and pituitary of the elasmobranch fish Scyliorhinus canicula, were studied by means of specific radioimmunoassay and immunohistochemistry using the indirect immunofluorescence method. Brain and pituitary extracts showed a good cross-reactivity with the ovine CRF antiserum, but serial dilutions of tissue samples did not completely parallel the standard curve. Relatively high concentrations of CRF-like material were found within the pituitary, diencephalon, and telencephalon. CRF-like immunoreactive perikarya were observed in the preoptic nucleus and in the nucleus lateralis tuberis. Numerous immunoreactive cells appeared to be of the CSF-contacting type. CRF-like immunopositive fibers were seen to run through the hypothalamus within the ventro-medial floor of the infundibular region. A dense plexus of immunoreactive nerve endings terminated in the median eminence and the neurointermediate lobe of the pituitary. These results indicate that a neurosecretory system containing CRF-like immunoreactivity exists in the brain of elasmobranchs, a group of vertebrates which has diverged early from the evolutionary line leading to mammals. In addition, our data support the notion that a CRF-like molecule is involved in the regulation of corticotropic and melanotropic cell activity in this primitive species of fish.     

ACTH-releasing activity of urotensin I and ovine CRF: interactions with arginine vasotocin, isotocin and arginine vasopressin

The release of ACTH from superfused dispersed goldfish anterior pituitary cells was examined to determine if the neurohypophyseal peptides arginine vasotocin (AVT), isotocin (IST) or arginine vasopressin (AVP) potentiate the ACTH-releasing activities of the structurally homologous peptides urotensin I (UI) or ovine corticotropin-releasing factor (CRF). The ACTH-releasing activities of the neurohypophyseal peptides and UI or CRF were additive. AVT, IST or AVP failed to potentiate the ACTH-releasing activity of UI or CRF. These results suggest that in teleost fishes neurohypophyseal peptides have intrinsic ACTH-releasing activity but, unlike mammals, do not potentiate the release of ACTH evoked by CRF, or by the piscian CRF-like peptide, UI.     

Immunocytochemical localization of vasotocin-like immunoreactivity in the brain of the cartilaginous fish,Scyliorhinus caniculus

Summary The distribution of vasotocin-like peptides in the central nervous system of the cartilaginous fish Scyliorhinus canicula was determined by indirect immunofluorescence and peroxidase anti-peroxidase techniques, using a specific antiserum raised in rabbits against synthetic vasotocin. Immunoreactive perikarya were mainly detected in the anterior hypothalamus, within the midcaudal part of the preoptic nucleus. The most rostral positive cell bodies were located in the dorso-lateral parts of the preoptic area, whereas at a more caudal level, they took a ventro-medial position within the deepest layers of the nucleus. Throughout the preoptic region these cells varied in shape according to their location. Occasionally, scattered vasotocin-like immunopositive cells were also identified in the nucleus periventricularis hypothalami. Vasotocin immunoreactivity was detected in numerous varicose nerve fibers of the preopticohypophysial tract. These fibers were seen to course through the medio-basal hypothalamus and caudally, after having passed the hypophysial stem, they reached the neurointermediate lobe of the pituitary. Numerous immunoreactive fibers were also observed within the rostro-medial region of the median eminence. At this level the fibers were in close proximity to the capillary loops. In the preoptic region, some stained cells exibited short processes that appeared to contact non-reactive perikarya. By comparing the distribution of vasotocin- and corticotropin-releasing factor immunoreactivity on adjacent then serial sections, it was revealed that these peptides, in S. canicula, do not coexist in the same perikarya. The present results, are compared with those obtained in other vertebrate groups, and their possible functional implications are discussed.     

Immunocytochemical localization and spatial relation to the adenohypophysis of a somatostatin-like and a corticotropin-releasing factor-like peptide in the brain of four amphibian species

Summary The distribution of somatostatin (SRIF) — and corticotropin-releasing factor (CRF)-like — immunoreactive material was studied in the brain of four amphibian species (Ambystoma mexicanum, Pleurodeles waltlii, Xenopus laevis, Rana ridibunda) by use of immunocytochemistry. A wide network of SRIF-immunoreactive fibers and numerous perikarya were observed in all amphibians examined, with a dense accumulation of nerve endings in the external layer of the median eminence (ELME). In the representatives of the four amphibian species the CRF-like system was more circumscribed. Immunoreactive perikarya were present in the preoptic area, mainly in a ventrobasal position, and in the interpeduncular nucleus. The tract running along the ventral part of the tuber cinereum ends in the ELME facing the rostroventral lobe of the pars distalis that contains corticotrophs. CRF fibers were scarce or absent in the neural lobe. In all species studied in the present work, CRF fibers end in the area of the ELME close to the pituitary lobe containing corticotrophs. This correlation is similar to that reported for the Japanese quail and several teleosts.This work was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek and the CNRS     

Immimohistochemical localization of corticotropin-releasing factor in selected brain areas of the European starling (Sturnus vulgaris) and the song sparrow (Melospiza melodia)

Summary Corticotropin-releasing factor (CRF) was localized in the brains of two passerine species, the European starling (Sturnus vulgaris) and the song sparrow (Melospiza melodia), by means of immunohistochemistry. The hypothalamic distribution of this peptide in these species includes a complex of immunoreactive perikarya observed in the paraventricular nucleus (PVN), in both its medial and lateral divisions. Nerve fibers were also seen running from these areas to the anterior median eminence (AME) where a terminal field is apparent. A wide variety of extra-hypothalamic nuclei containing CRF-immunoreactive cells and fibers were identified. An apparent CRF terminal field can be visualized in the lateral septum. A dense fiber plexus is present in the nucleus accumbens (Ac) and more caudally in the nucleus of the stria terminalis (nST). In colchicinepretreated animals, it was revealed that these areas also contain CRF-stained perikarya. The pattern of CRF immunoreactivity in the Ac-nST complex is continuous, with no distinction apparent between the nuclei. The medial preoptic area (mPOA) and the adjacent diagonal band of Broca contain CRF-fibers, while cells are apparent in the mPOA. In the mesencephalon, cells were visualized in the midbrain central gray; a terminal field and scattered positively stained perikarya were found in areas more ventral to the central grey that are adjacent to the third cranial nerve. Scattered cells were also seen at the border of the nucleus intercollicularis-nucleus mesencephalicus lateralis, pars dorsalis complex. In contrast to mammalian studies, no immunoreactive nerve fibers or perikarya were observed in telencephalic areas homologous to the mammalian neocortex. These studies confirm the presence of a CRF path-way regulating pituitary function and suggest a broad role played by CRF as a neuromodulator or neurotransmitter in autonomic and possibly behavioral activities in these species.     

The regulation of the corticomelanotropic cell activity in Aves--II. Effect of various peptides on the release of ACTH from dispersed, perfused duck pituitary cells

We have estimated the corticotropin-releasing activity (CRA) of different neurohypophyseal peptides and synthetic corticotropin-releasing factor (CRF) in the duck, using perfused dispersed pituitary cells and an ACTH radioimmunoassay adapted to duck material. Log dose-response curves were obtained for different doses of arginine-vasopressin (AVP), arginine-vasotocin (AVT), mesotocin (MT), oxitocin (OT) and ovine CRF (oCRF) and compared to the response obtained with dilutions of duck median eminence extracts (DME). All peptides tested behaved as partial agonists compared to DME. AVT and MT were the most potent of all peptides tested, with a capacity of 60% relative to DME. CRF was a weak agonist together with AVP and OT. AVT and CRF perfused together at equal doses significantly potentiated the effect of each other, yielding a dose-response line whose slope approximated that of DME. A similar design was used to test the CRA of the same substances in the rat. The main difference in the pattern of response between the two species was the low potency displayed by all the neurohypophyseal peptides in the rat, compared with CRF which, in contrast with what occurred with the duck system, was the most potent secretagogue of all peptides tested. It is concluded that in birds, as in mammals, the control of ACTH secretion may be exerted by neurohypophyseal peptides and a CRF-like peptide acting synergistically upon the corticomelanotropic cell.     

Comparative distribution of urocortin- and CRF-like immunoreactivities in the nervous system of the earthworm Lumbricus terrestris

Corticotropin-releasing factor (CRF) and urocortin (Ucn) are both members of the CRF neuropeptide family. The distribution of Ucn- and CRF-like immunoreactive (ir) structures in the central nervous system of several vertebrate species has been studied, but little is known about that in non-vertebrates. We used a highly specific polyclonal antibody against rat Ucn and CRF to determine and compare the distribution of Ucn- and CRF-like immunoreactivity in the earthworm nervous system. Several Ucn- and CRF-like ir perikarya were described in the cerebral ganglion, subesophageal and ventral cord ganglia. The majority of Ucn-like ir cells were found in the ventral ganglia, whereas CRF-like ir cells were most abundant in the cerebral ganglion. Scattered Ucn- and CRF-like ir varicose fiber terminals were seen in all areas of the earthworm central nervous system. Ucn-like ir cell bodies and fiber terminals were also demonstrated in the pharyngeal wall. No co-localization of Ucn- and CRF-like ir nervous structures were observed. This study provided morphological evidence that Ucn- and CRF-like neurosecretory products exist in the earthworm central nervous system. Furthermore, both the distribution and morphology of Ucn- and CRF-like ir structures were distinct, therefore, it can be hypothesized that these neuropeptides exert different neurendocrine functions in the earthworm nervous system.     

Localization of growth hormone-releasing factor-like immunoreactivity in the hypothalamo-hypophysial system of some teleost species

Summary An antiserum to growth hormone-releasing factor (GRF) 1-44 was applied on brain and pituitary sections of nine teleost species. Immunoreactive (ir) perikarya were demonstrated in parvo- and magnocellular portions of the preoptic nucleus (PON) and occasionally in the nucleus lateralis tuberis. The two tracts originating in the PON ran ventro-laterally toward the optic chiasm and then caudally in the basal hypothalamus. In the pars distalis (PD) of the eel, carp, goldfish and salmonids, GRF-ir fibers did not enter the rostral PD and few fibers passed close to somatotropes. In.Myoxocephalus andMugil, a variable number of ir-fibers passed close to cells of the rostral and proximal PD. In the neurointermediate lobe, GRF-ir fibers were located exclusively in the neural tissue of the eel and trout. In goldfish, carp andMyoxocephalus, GRF-ir fibers entered the intermediate lobe. This antiserum also labeled corticotrops and, to a lesser extent, melanotrops in the pituitary of cyprinids. A variable number of perikarya contained both GRF and vasotocin in the PON of the eel. In all teleost species studied so far, the distribution patterns of GRF are different, and the function of the various adenohypophysial cell types appears to be differently modulated, according to the variable distribution of GRF in the pituitary.     

Distribution of NADPH-diaphorase and nitric oxide synthase in the trigeminal ganglion and mesencephalic trigeminal nucleus of the cat. A histochemical and immunohistochemical study

The trigeminal ganglion (TrG) and mesencephalic trigeminal nucleus (MTN) neurons are involved in the transmission of orofacial sensory information. The presence of nitric oxide (NO), a putative neurotransmitter substance in the nervous system, was examined in the cat TrG and MTN using nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry and nitric oxide synthase (NOS) immunohistochemistry. In the TrG, where the majority of the trigeminal primary afferent perikarya are located, most of the intensely NADPH-d/ NOS-stained cells were small in size and distributed randomly throughout the ganglion. The medium-sized neurons were moderately stained. A plexus of pericellular varicose arborizations around large unstained ganglion cells and densely stained fibers in-between could also be observed. In the caudal part of the MTN, both NADPH-d activity and NOS immunoreactivity was present in MTN neurons. In addition, a few scattered NADPH-d/NOS-containing neurons were found in the mesencephalic-pontine junction part of the nucleus. In contrast, only nerve fibers and their terminals were present at a more rostral level in the mid- and rostral MTN. MTN neuronal perikarya were enveloped in fine basket-like NADPH-d/ NOS-positive networks. Differential expression patterns of NOS and its marker NADPH-d suggest that trigeminal sensory information processing in the cat MTN is controlled by nitrergic input through different mechanisms. We introduce the concept that NO can act as a neurotransmitter in mediating nociceptive and proprioceptive information from periodontal mechanoreceptors but may also participate in modulating the activity of jaw-closing muscle afferent MTN neurons.     

The hypothalamo-hypophyseal system of the white seabream Diplodus sargus: immunocytochemical identification of arginine-vasotocin,isotocin, melanin-concentrating hormone and corticotropin-releasing factor

The distribution of the neurosecretory hormones vasotocin, isotocin and melanin-concentrating hormone and the hypophysiotropic hormone corticotropin-releasing factor was studied in the hypothalamo-hypophyseal system of the white seabream (Diplodus sargus) using immunocytochemical techniques. Magnocellular and parvocellular perikarya immunoreactive for arginine-vasotocin and isotocin were present in the nucleus preopticus. Perikarya immunoreactive for arginine-vasotocin extended more caudally with respect to isotocin-immunoreactive perikarya. Parvocellular perikarya were located at rostroventral levels and magnocellular perikarya in the dorsocaudal portion of the nucleus. Arginine-vasotocin and isotocin did not coexist in the same neuron. Fibres immunoreactive for arginine-vasotocin and isotocin innervated all areas of neurohypophysis and terminate close to corticotropic and melanotropic cells. Perikarya immunoreactive for melanin-concentrating hormone and corticotropin-releasing factor were observed in the nucleus lateralis tuberis, with a few neurons in the nucleus periventricularis posterior. In addition, melanin-concentrating hormone immunoreactive perikarya were detected in the nucleus recessus lateralis. The preoptic nucleus did not show immunoreactivity for these antisera. Fibres showing melanin-concentrating hormone and corticotropin-releasing factor immunoreactivity ended close to the melanotropic and somatolactotrophic cells of the pars intermedia, and close to the corticotrophic cells of the rostral pars distalis.     

Immunohistochemical localization of corticotropin-releasing factor (CRF)-containing neurons in the hypothalamus of the Japanese quail,Coturnix coturnix

Immunohistochemical localization of corticotropin-releasing factor (CRF)-like immunoreactivity in the brain of the Japanese quail was studied by means of the peroxidase anti-peroxidase (PAP) method. CRF-immunopositive perikarya of parvocellular neurons were observed mainly in the nucleus praeopticus medialis and nucleus paraventricularis. Additional perikarya were also detected in the nucleus hypothalamicus posterior medialis in the hypothalamus and in the non-hypothalamic nucleus accumbens, nucleus septalis lateralis and nucleus dorsomedialis and dorsolateralis thalami. No CRF immunoreaction was found to coexist with the vasotocin (Vt)-containing system in comparative examination of consecutive sections treated with anti-vasopressin (Vp) serum. The CRF-immunoreactive fibers were detected mainly in the external layer of the anterior median eminence but not in its posterior division. Unilateral adrenalectomy induced the marked reduction in number of the CRF immunopositive fibers in the anterior median eminence.     

Immunocytochemical localization of a galanin-like peptidergic system in the brain and pituitary of some teleost fish.

Immunostaining of brain and pituitary sections of teleost fishes (eels, salmonidae, cyprinidae, gourami, sculpin, mullet) with anti porcine galanin (GAL) revealed the presence of immunoreactive (ir) perikarya and a rich network of fibers. Ir-perikarya were located rostrodorsally to the recessus preopticus, and in the posterior tuberal hypothalamus. Ir-fibers were abundant in basal telencephalon and around diencephalic ventricular recesses but never contacted their lumen. Furthermore, they were observed in basal hypothalamus, brainstem and ventral medulla. Ir-fibers passed along corticotropic (ACTH), gonadotropic cells and somatotropes (GH cells) in eel and trout pars distalis, but rarely ended in caudal neurohypophysis. In goldfish pituitary ir-fibers occurred in neural digitations and among different cell types which however did not contain a GAL-like peptide. The relation GAL fibers/GH cells appeared more evident in species with a high growth rate. The other species showed a similar distribution of brain GAL. In eels and trout, ir-perikarya were not observed in areas containing somatostatin, GH- and ACTH-releasing factor, and ACTH-like perikarya, suggesting that GAL did not coexist with these peptides. The widespread distribution of a GAL-like peptide in teleost brain suggests that it could play a role of neurotransmitter and/or neuromodulator and regulate the secretion of adenohypophysial hormone(s).     

Galanin-like immunoreactivity is increased in the brain of estradiol- and methyltestosterone-treated eels.

A galanin-like peptidergic system was demonstrated in the brain of Anguilla. A group of immunoreactive perikarya was located in the nucleus preopticus periventricularis close to the recessus preopticus. Galaninergic fibers occurred in various brain areas. Galanin identified in mammalian pituitary cells was undetectable in fish adenohypophysial cells. Estradiol increased the immunostaining of the rostral perikarya and brain fibers in both male and female European eels kept in fresh water and in female American eels in sea water. Methyltestosterone, an aromatizable androgen, increased galanin immunoreactivity in rostral perikarya and brain fibers of male European eels and female American eels. The cross-sectional area of these perikarya increased significantly after both treatments whereas cell bodies of the posteroventral hypothalamus were slightly affected. Dihydrotestosterone showed no clear effect. Fibers close to the corticotropes were sometime increased, but galanin synthesis was not induced in pituitary cells. In contrast, estradiol induced galanin synthesis in rat pituitary cells, but had a still controversed effect on hypothalamic galanin. A putative influence of galanin on the pituitary-gonadal axis is discussed as gonadal hormones diversely affect gonadotropes and gonosomatic indices in Anguilla.     

Immunoreactive beta-endorphin and ACTH in the same neurons of the hypothalamic arcuate nucleus in the rat.

Immunoreactive ACTH and beta-endorphin (beta-End) were localized in the brain and pituitaries of normal and colchicine-treated rats, using the immunoperoxidase method at the light microscopic level. On adjacent serial 5-micron paraffin sections of anterior pituitaries, both ACTH and beta-End could be found in the same cells. On adjacent 5-micron paraffin sections of brains of colchicine-treated rats, both ACTH and beta-End could be found in the same perikarya of hypothalamic arcuate nucleus neurons. It appeared that all perikarya containing beta-End contained ACTH as well, suggesting that neurons producing beta-End also produce ACTH. Pathways of ACTH fibers corresponded to pathways of beta-End fibers. These findings suggest that the synthesis, and transport, of ACTH and beta-End are linked in the brain as well as in the pituitary, possibly through a common precursor.