Human kisspeptins activate neuropeptide FF2 receptor.

We studied the possible activation of a neuropeptide FF2 receptor (NPFF2R) by kisspeptins, neuropeptides derived from the mouse and human metastin or Kiss-1 precursor. The hypothesis was that the human kisspeptins, which share the C-terminal dipeptide RF-NH(2) with NPFF, might activate the NPFF2R, as has previously been shown for two related peptides, prolactin-releasing peptide and RF-amide-related peptide. Using two-electrode voltage clamp of Xenopus oocytes, we found that 100 nM NPFF strongly activated the human NPFF2R expressed together with rat GIRK1/4 inward rectifier potassium channels, and that 100 nM hKisspeptin-13 and hKisspeptin-8 had about 25% relative efficacy to that of NPFF. The current response induced by hKisspeptin-13 was proportional to its concentration (1-500 nM). The corresponding mouse peptides resulted in low activation only. When hNPFF2R was expressed in Chinese hamster ovary (CHO) cells, NPFF and its stable analog (1DMe)Y8Fa induced guanosine 5'-(gamma-[(35)S]thio)-triphosphate (GTP-gamma-[(35)S]) binding with EC(50) values of 13+/-4 and 16+/-4 nM, respectively. hKisspeptin-13 induced the binding with an EC(50) value of 110+/-50 nM, whereas mKisspeptin-13 induced very modestly activation with an EC(50) value>2 microM. The results suggest that, besides regulation of reproduction, kisspeptins have a potential to mediate physiological effects on, for example autonomic regulation and nociception in man via the NPFF2R pathways.

Mouse mammary epithelial histamine system.

Histamine is suggested to play a role in mammary gland growth regulation, differentiation and functioning during pregnancy and lactation. Two pools of histamine are thought to be involved in these processes: mastocyte- and epithelial cell related histamine. In the present study we focused on epithelial cells. Immunohistochemistry has shown that the epithelial cells positive for histamine and L-histidine decarboxylase (HDC), the primary enzyme regulating histamine biosynthesis, were mainly found in cells forming alveolar structures in the mammary gland. Cultured primary mouse mammary epithelial cells (MMEC) expressed strong HDC immunoreactivity, especially dividing cells and non-differentiated ones. Histidine decarboxylase activity undergoes significant changes during pregnancy and lactation. Pregnancy associated intensive growth of the mammary gland coincided with an increase and the first days of lactation with a decrease of HDC protein expression. Binding studies with mammary tissue membranes and epithelial cell membranes revealed the presence of H1 and H3 but not H2 receptors. Summarizing, our data have shown that mammary epithelial cells are capable of synthesizing and excreting histamine and they bear histamine receptors. These findings further substantiate the role of histamine in mammary gland physiology.

Histamine in brain development and tumors.

Histamine is found in developing mammalian brain in both neurons and mast cells. Under normal conditions, histamine H1 and H2 receptors are found in neural, glial and endothelial cells, and H3 receptors at least on neurons. Experimental brain tumors display both H1 and H2 receptors, and histamine increases permeability in the tumors and in the neighboring areas. Many studies have addressed histaminergic signalling mechanisms in cell lines originating from brain tumors. However, the role of histamine in normal development of brain structures, proliferation and differentiation of neurons and glial cells, and growth of malignant tumors in situ is still poorly understood.

Estrogen receptor immunoreactivity is present in the majority of central histaminergic neurons: evidence for a new neuroendocrine pathway associated with luteinizing hormone-releasing hormone-synthesizing neurons in rats and humans.

The central regulation of the preovulatory LH surge requires a complex sequence of interactions between neuronal systems that impinge on LH-releasing hormone (LHRH)-synthesizing neurons. The reported absence of estrogen receptors (ERs) in LHRH neurons indicates that estrogen-receptive neurons that are afferent to LHRH neurons are involved in mediating the effects of this steroid. We now present evidence indicating that central histaminergic neurons, exclusively located in the tuberomammillary complex of the caudal diencephalon, serve as an important relay in this system. Evaluation of this system revealed that 76% of histamine-synthesising neurons display ERalpha-immunoreactivity in their nucleus; furthermore histaminergic axons exhibit axo-dendritic and axo-somatic appositions onto LHRH neurons in both the rodent and the human brain. Our in vivo studies show that the intracerebroventricular administration of the histamine-1 (H1) receptor antagonist, mepyramine, but not the H2 receptor antagonist, ranitidine, can block the LH surge in ovariectomized estrogen-treated rats. These data are consistent with the hypothesis that the positive feedback effect of estrogen in the induction of the LH surge involves estrogen-receptive histamine-containing neurons in the tuberomammillary nucleus that relay the steroid signal to LHRH neurons via H1 receptors.

Association between myocardial infarction and the mast cells in the adventitia of the infarct-related coronary artery.

BACKGROUND: Histamine, a product of mast cells, is an effective vasoconstrictor of atherosclerotic coronary arteries. Because it has been suggested that coronary spasm plays a role in acute coronary syndromes such as myocardial infarction (MI), we quantified and characterized the mast cells in the adventitia of infarct-related coronary arteries. METHODS AND RESULTS: In a series of 17 autopsied MI patients, we identified the segment of the left coronary artery with ruptured plaque responsible for the infarction. More distal segments from the infarct-related coronary artery, either with nonruptured plaques or with normal intima, were also studied. Corresponding segments taken from left coronary arteries obtained from 17 patients who had died of noncardiac causes served as controls. Adventitial mast cells in the infarct-related and the control coronary arteries were identified immunohistochemically by staining for tryptase. In the infarct-related coronary arteries, we also stained for chymase and histamine. Moreover, T lymphocytes and macrophages were identified immunohistochemically and counted. In the infarct-related coronary arteries, significantly larger numbers of mast cells were present in the adventitia backing ruptured plaques (98+/-40 mast cells/mm2, mean+/-SD) than in the adventitia backing nonruptured plaques (41+/-12 mast cells/mm2; P<0.001) or backing normal intima (19+/-8 mast cells/mm2; P<0.001). No such difference was found among the 3 different segments in the control coronary arteries. The majority of mast cells contained not only tryptase but also chymase. Mast cells were the only cells in the coronary adventitia that contained histamine. The proportion of adventitial mast cells that were degranulated was highest in the segments with ruptured plaques. The numbers of adventitial macrophages and T lymphocytes were also increased in the segments with plaque rupture. CONCLUSIONS: In infarct-related coronary arteries, the number of degranulated mast cells in the adventitia backing ruptured plaques is increased. Histamine released from the degranulated mast cells may reach the media, where it may locally provoke coronary spasm and thus contribute to the onset of MI.

ECL cell morphology.

Using immunohistochemistry at the conventional light, confocal and electron microscopic levels, we have demonstrated that rat stomach ECL cells store histamine and pancreastatin in granules and secretory vesicles, while histidine decarboxylase occurs in the cytosol. Furthermore the ECL cells display immunoreactivity for vesicular monoamine transporter type 2 (VMAT-2), synaptophysin, synaptotagmin III, vesicle-associated membrane protein-2, cysteine string protein, synaptosomal-associated protein of 25 kDa, syntaxin and Munc-18. Using electron microscopy in combination with stereological methods, we have evidence to suggest the existence of both an exocytotic and a crinophagic pathway in the ECL cells. The process of exocytosis in the ECL cells seems to involve a class of proteins that promote or participate in the fusion between the granule/vesicle membrane and the plasma membrane. The granules take up histamine by VMAT-2 from the cytosol during transport from the Golgi zone to the more peripheral parts of the cells. As a result, they turn into secretory vesicles. As a consequence of stimulation (e.g., by gastrin), the secretory vesicles fuse with the cell membrane to release their contents by exocytosis. The crinophagic pathway was studied in hypergastrinemic rats. In the ECL cells of such animals, the secretory vesicles were found to fuse not only with the cell membrane but also with each other to form vacuoles. Subsequent lysosomal degradation of the vacuoles and their contents resulted in the development of lipofuscin bodies.

Histamine in the chick pineal gland: origin, metabolism, and effects on the pineal function.

The chick pineal gland contains histamine and tele-methylhistamine. The levels of both substances are elevated after treatment of chicks with the amino acid precursor of histamine, L-histidine (1 g/kg, ip). In control and L-histidine-loaded animals the pineal levels of histamine and tele-methylhistamine are higher in light-exposed than in dark-adapted animals (measured at the end of the light phase and in the middle of the dark phase of 12 hr light, 12 hr dark illumination cycle, respectively). The chick pineal gland contains histamine-immunofluorescent cells displaying mast cell morphology; they are seen in the vicinity of the capsule and in the parenchyma. Enzymatic studies showed the presence of the activity of histamine synthesizing and inactivating enzyme, i.e., L-histidine decarboxylase (HDC) and histamine-methyltransferase (HMT). The detected enzyme activities were sensitive to specific inhibitors of HDC (alpha-fluoromethylhistidine and alpha-hydrazinohistidine) and HMT (quinacrine and metoprine); inhibitors of aromatic amino acid decarboxylase alpha-methyl-DOPA and NSD-1015 were inactive on HDC. Exogenous histamine added to organ-cultured chick pineals strongly stimulated endogenous cyclic AMP accumulation and moderately increased melatonin secretion. The data, considered collectively, suggest that in avians histamine, probably originating from the pineal mast cell compartment, may function as a regulator of pineal gland activity.

Expression and intracellular localization of catechol O-methyltransferase in transfected mammalian cells.

The intracellular localization of soluble and membrane-bound isoforms of rat and human catechol O-methyltransferase (COMT) was studied by expressing the recombinant COMT proteins either separately or together in mammalian cell lines (HeLa and COS-7 cells) and in rat primary neurons. The distribution of soluble and membrane-bound COMT enzyme was visualized by immunocytochemistry. For comparison, the localization of native COMT was studied in rat C6 glioma cells by immunoelectron microscopy. Staining of cells expressing membrane-bound COMT with a COMT-specific antiserum revealed an immunofluorescence signal in intracellular reticular structures and in the nuclear membrane. Double-staining of the cells with antisera against proteins specific for the rough endoplasmic reticulum indicated that they colocalized with membrane-bound COMT, suggesting that it resided in the endoplasmic reticulum. Notably, no COMT-specific fluorescence of plasma membranes was detected. The signal in the endoplasmic reticulum was also evident in the cells expressing both recombinant COMT forms. Intracellular native COMT reaction was detected by immunoelectron microscopy in rat C6 glioma cells and an intense cytoplasmic signal was seen in the primary neurons infected with the recombinant Semliki Forest virus. The cells expressing recombinant soluble COMT revealed intense nuclear staining together with diffuse cytoplasmic immunoreactivity, suggesting that a part of soluble COMT is transported to nuclei. Western blotting from rat liver and brain revealed soluble COMT in the nuclei. Enzyme activity measurements from liver cytoplasmic and nuclear fractions suggested that about 5% of the soluble COMT resided in nuclei. The intracellular localization of both COMT forms implies that COMT acts in the cytoplasm and possibly also in the nuclear compartment, and that the physiological substrates of COMT enzymes may have to be internalized before their methylation by COMT.

Gastric mucosal histamine storing cells. Evidence for different roles of mast cells and enterochromaffin-like cells in humans.

Gastric mucosal histamine content, enterochromaffin-like cell density, and mast cell density were studied in 13 subjects under omeprazole therapy, 13 partially gastrectomized subjects with a Billroth II reconstruction, 10 partially gastrectomized subjects with a Roux-en-Y reconstruction, and 9 control subjects. Histamine content was significantly greater both in the subjects with higher gastrinemic levels (omeprazole-treated subjects) and those with more abundant enterogastric reflux (Billroth II subjects) than in controls. Enterochromaffin-like cell density was significantly greater in the omeprazole subjects than in each of the other groups. Mast cell density was significantly greater in Billroth II subjects than in controls. Serum gastrin levels, mucosal histamine content, and enterochromaffin-like cell density were positively correlated. Gastrin was not correlated to mast cell density. These results support the existence of different control pathways for enterochromaffin-like and mast cells. Moreover, they suggest that enterochromaffin-like cells and mast cells are involved in the regulation of gastric secretion and in gastric mucosal injury-repair mechanisms, respectively, due to histamine release.

Developmental patterns of histamine-like immunoreactivity in the mouse.

We studied the appearance and distribution of histamine (HA) during mouse embryogenesis, neonatal period, and adulthood using a specific rabbit HA antiserum and indirect immunofluorescence. HA first appeared on the Embryonic Day 13 (E13) in scattered mast cells in the gastrointestinal (GI) muscularis externa and liver. The splenic primordium contained a dense population of intensely HA-immunoreactive (HA-ir) cells from E13 on. From E15 to the birth, HA was detected in many embryonic cell types. On E15, the first HA-ir epithelial endocrine cells appeared in the oxyntic mucosa. In addition to the HA-ir cells in GI tract and liver, some nerve cells in ganglia of the peripheral nervous system (PNS), some fibers in spinal and cranial nerves, nerve fibers in mesenterium, and nerve plexuses of the gastrointestinal muscularis externa were HA-ir from E15 on. Occasional HA-ir nerve fibers were detected within the glandular epithelium of the oxyntic mucosa, pancreas, and salivary glands during late embryogenesis. During the same period, bright fluorescence was observed in cells of the kidney convoluted tubules and pancreatic islet cells. From E14 on, mast cells exhibiting bright fluorescence were scattered throughout the connective tissue of the fetus, and their number increased rapidly with age. Their density was especially high in subcutaneous connective tissue. Embryonic epidermal cells showed faint HA immunoreactivity. In musculoskeletal tissues, developing bone and occasional striated muscle cells exhibited HA immunoreactivity. Interestingly, most cells in liver showed transiently weak HA immunoreactivity during embryogenesis. In adult mouse, HA was stored only by scattered mast cells, oxyntic epithelial cells, and neurons in the tuberomamillary nucleus of the brain. The other HA-containing embryonic cells were negative for HA in adult mouse. In conclusion, HA immunoreactivity is widely distributed in epithelial, neuronal, and mast cells in various organs during mouse embryogenesis.

Endogenous histamine in cultured bovine adrenal chromaffin cells.

Histamine releases catecholamines and opioids in primary cultured bovine adrenal medullary (BAM) chromaffin cells. We have studied whether histamine is synthesized and localized in BAM cells, and whether it can be released upon activation with secretagogues. In BAM cells histamine is immunohistochemically co-localized with tyrosine hydroxylase in 45 +/- 8% of all cells. Only histamine immunoreactivity was observed in 8 +/- 2% of all BAM cells. No mast-cell-like cells were observed in our system. Histamine can be released from BAM cells by high potassium (56 mM K+) in a calcium-dependent manner. Compound 48/80 did not release histamine from BAM cells but nicotine caused a dose-dependent liberation of the amine. Cultured BAM cells have histidine decarboxylase activity which is inhibited by alpha-fluoromethylhistidine. These results indicate that endogenous histamine is synthesized, stored and released in BAM chromaffin cells in vitro.

Multiple neurotransmitters in the tuberomammillary nucleus: comparison of rat, mouse, and guinea pig.

Tuberomammillary neurons in the posterior hypothalamus are the sole source of neuronal histamine in adult mammalian brain. In the rat, these cells are reported to contain immunoreactivity for gamma-aminobutyric acid (GABA) and several neuropeptides. We compared the presence of these substances in the tuberomammillary cells of the rat, mouse, and guinea pig. In all three species, all histamine-immunoreactive neuronal cell bodies were positive for GABA. This suggests that GABAergic transmission may be important in tuberomammillary function. No cell bodies immunoreactive for thyrotropin releasing hormone (TRH) were found in the guinea pig or mouse tuberomammillary area. In contrast, about 14% of the histamine-immunoreactive tuberomammillary cells in the rat were TRH-positive. These cells were small or medium-sized and were located only in the medial part of the tuberomammillary complex. An antibody against porcine galanin stained about 45% of the tuberomammillary cell bodies in the rat and about 28% in the mouse, but none in the guinea pig. A large proportion of the cells in the rat and mouse, but none in the guinea pig, were positive for met-enkephalin-arg-phe. In contrast, all histamine-containing tuberomammillary cells in the guinea pig, but none in the rat or mouse, were immunoreactive for met-enkephalin. This may indicate a different expression of proenkephalin-derived peptides in the tuberomammillary neurons in these species. Some substance P-immunoreactive cell bodies were located in the tuberomammillary area in all three species. However, only 3% of the histamine-immunoreactive cell bodies in the rat and mouse but none in the guinea pig were substance P-positive. The neurochemical properties of the tuberomammillary nucleus that exhibited species commonality deserve to be studied neurochemically and electrophysiologically in order to determine the functional relevance of coexisting transmitters in this nucleus.

Distribution of taurine in the rat cerebellum and insect brain: application of a new antiserum against carbodiimide-conjugated taurine.

The production, specificity and application of an antiserum against taurine conjugated to succinylated ovalbumin by means of 1-ethyl-3(3-dimethylaminopropyl)-carbodiimide is reported. The antiserum was produced in rabbits. The carbodiimide was used also as a tissue fixative. The development of the antibody titre was followed by dot-blot tests on nitrocellulose filters using different amino acid conjugates and with immunohistochemical reaction in the rat and insect brain. Blocking controls were also used. Taurine antiserum, sufficiently specific and sensitive, developed after the fourth booster injection, after which the antiserum was characterized. In the insect brain, intense taurine-like immunoreactivity was observed in the photoreceptors, in the Kenyon cells and the neuropile of the mushroom bodies, in the lower part of the central body and in the antennal lobes. In the rat carebellum, intense taurine-like immunoreactivity was seen in the Purkinje cells. Immunoreaction was seen also in small cells most probably corresponding to the basket cells. The use of the carbodiimide in the production of antisera against taurine provides a parallel method for comparison of the distribution of taurine-like immunoreactivity obtained with antisera made against conjugates prepared with aldehydes.

Helodermin- and helospectin-like immunoreactivities in the rat brain: an immunochemical and immunohistochemical study.

Helodermin is an amidated peptide of 35 amino acid residues isolated from the lizard Heloderma suspectum. Homologous peptides, helospectins I and II, peptides of 38 and 37 amino acid residues, respectively, have been isolated from the lizard Heloderma horridum. This group of peptides stimulates the adenylate cyclase activity. Helodermin- and helospectin-like immunoreactivities were studied in the rat brain by using immunohistochemistry and radioimmunoassay in combination with high-performance liquid chromatography. The highest concentrations of helodermin-like immunoreactivity were found in the cerebellum and hypothalamus. The chromatographic analysis of rat brain extract revealed one main immunoreactive peak with elution properties similar to those of authentic lizard helodermin. Helodermin-immunoreactive neurons were located in the supraoptic nucleus, suprachiasmatic nucleus, periventricular nucleus, arcuate nucleus and central gray. Fibers and terminals of varying densities were observed in the bed nucleus of the stria terminalis, medial part of the central nucleus of amygdala, external layer of the median eminence, thalamus and central gray. The highest concentrations of helospectin-like immunoreactivity were found in the cerebral cortex, hypothalamus and medulla. The chromatographic analysis of brain extract revealed one major peak with elution properties similar to those of authentic helospectin I. Helospectin-immunoreactive neurons were located in the suprachiasmatic nucleus, central gray, cerebral cortex, dorsomedial hypothalamic nucleus and supramammillary nucleus. Helospectin-immunoreactive fibers and terminals were found in the bed nucleus of the stria terminalis, medial part of the central nucleus of amygdala, median eminence, lateral parabrachial nucleus, central gray, cerebral cortex, thalamus and nucleus of the solitary tract. The present study has revealed novel neuronal systems in the rat brain by using antisera against the lizard peptides helodermin and helospectin. The patterns of immunostaining suggest a role for the helodermin- and helospectin-like peptides in the hypothalamo-hypophyseal control of endocrine functions.

The distribution of pituitary adenylate cyclase-activating polypeptide-like immunoreactivity is distinct from helodermin- and helospectin-like immunoreactivities in the rat brain.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is an amidated 38-residue polypeptide isolated from the ovine hypothalamus. Helodermin, a 35-amino acid peptide, and helospectins, peptides of 38 and 37 amino acid residues, have been isolated from lizard venom. PACAP, helodermin and helospectins share structural features and have a similar profile of pharmacological effects: they stimulate adenylate cyclase. We studied the distribution and characteristics of PACAP-like immunoreactivity in the rat brain with immunochemical and immunohistochemical methods and compared its distribution with that of helodermin- and helospectin-like immunoreactivities. With radioimmunoassay, the highest concentrations of PACAP-like immunoreactivity were found in the hypothalamus and cerebellum. PACAP-immunoreactive cell bodies were located immunohistochemically in the supraoptic nucleus, paraventricular and periventricular hypothalamic nuclei, and in the central grey. PACAP-immunoreactive fibres and terminals were detected in the medial part of the central nucleus of amygdala, in the median eminence and neurohypophysis, and in the central grey. No PACAP-immunoreactive structures were observed in areas such as the cerebral cortex, hippocampus, or cerebellum. The distribution of PACAP-like immunoreactivity differed considerably from the distribution of helodermin- and helospectin-like immunoreactivities. The results of this study suggest that PACAP is a neuropeptide with a role in the regulation of endocrine function in the hypothalamo-hypophyseal axis.

Peptide YY-like immunoreactivity in sympathetic neurons of the rat.

The occurrence of peptide YY-like peptides in parts of the sympathetic nervous system of the rat was studied by immunocytochemistry and immunochemistry plus analysis by high performance liquid chromatography. Peptide YY-immunoreactive neurons and nerve fibers were detected in the superior cervical ganglion. Co-localization studies indicated that peptide YY and neuropeptide Y immunoreactivities co-exist in a subpopulation of neurons of the superior cervical ganglion. Immunochemical analysis revealed peptide YY-immunoreactive material, distinct from neuropeptide Y, in extracts of the superior cervical ganglion. On reverse-phase high performance liquid chromatography, extracts of superior cervical ganglion revealed several peaks of peptide YY-like immunoreactive material, one of which eluted close to the position of authentic porcine peptide YY. Peptide YY-immunoreactive nerve fibers were also present in sympathetic target tissues including the auricula and atria of the heart, carotid body, submandibular salivary gland and the adrenal cortex. It is suggested that peptide YY and/or peptide YY-like peptides are present not only in endocrine cells, but also in a subpopulation of cell bodies and fibers of the peripheral sympathetic nervous system.

The adhesive and neurite-promoting molecule p30: analysis of the amino-terminal sequence and production of antipeptide antibodies that detect p30 at the surface of neuroblastoma cells and of brain neurons.

A membrane-bound adhesive protein that promotes neurite outgrowth in brain neurons has been isolated from rat brain (Rauvala, H., and R. Pihlaskari. 1987. J. Biol. Chem. 262:16625-16635). The protein is an immunochemically distinct molecule with a subunit size of approximately 30 kD (p30). p30 is an abundant protein in perinatal rat brain, but its content decreases rapidly after birth. In the present study the amino-terminal sequence of p30 was determined by automated Edman degradations. A single amino-terminal sequence was found, which is not present in previously studied adhesive molecules. This unique sequence has a cluster of five positive charges within the first 11 amino acid residues: Gly-Lys-Gly-Asp-Pro-Lys-Lys-Pro-Arg-Gly-Lys. Antisynthetic peptide antibodies that recognize this sequence were produced in a rabbit, purified with a peptide affinity column, and shown to bind specifically to p30. The antipeptide antibodies were used, together with anti-p30 antibodies, to study the localization of p30 in brain cells and in neuroblastoma cells as follows. (a) Immunofluorescence and immunoelectron microscopy indicated that p30 is a component of neurons in mixed cultures of brain cells. The neurons and the neuroblastoma cells expressed p30 at their surface in the cell bodies and the neurites. In the neurites p30 was found especially in the adhesive distal tips of the processes. In addition the protein was detected in ribosomal particles and in intracellular membranes in a proportion of cells. (b) The antibodies immobilized on microtiter wells enhanced adhesion and neurite growth indicating that p30 is surface exposed in adhering neural cells. (c) Immunoblotting showed that p30 is extracted from suspended cells by heparin suggesting that a heparin-like structure is required for the binding of p30 to the neuronal cell surface. A model summarizing the suggested interactions of p30 in cell adhesion and neurite growth is presented.

Distribution of histamine in the cockroach brain and visual system: an immunocytochemical and biochemical study.

The distribution of histamine-immunoreactivity in the carbodiimide-fixed brain and visual system of the cockroach was revealed immunocytochemically with an antiserum against histamine (HA). Histamine levels were measured with high-pressure liquid chromatography. The results show a widespread distribution of histamine-containing somata and fibers in the brain, particularly in the visual system. The most intense immunolabeling was seen in the retinal photoreceptors and in the first optic ganglion, the lamina, where the short visual fibers make synaptic connections with the monopolar neurons, which also displayed immunofluorescence. Immunoreactive long visual fibers traversed the lamina and outer chiasma, terminating in the distal medulla. Tracts of histamine-immunopositive fibers appeared to link the optic ganglia to the protocerebrum. Prominent histamine-containing neurons were situated in the lateral protocerebrum. Immunolabeled pathways consisting of large-diameter fibers also were seen in the cockroach brain. The central parts of the brain, including the central body, were reached by thick immunoreactive fibers that gave rise to intensely fluorescent varicose processes there. In the mushroom bodies, immunoreactivity was limited to the calyces. The protocerebral bridge was nonreactive. Immunofluorescence was seen also in the antennal lobes, but not in the antennal nerves. The biochemical measurements correlated well with the immunocytochemical data. The retinas and optic lobes, measured together, contained remarkably large amounts of histamine. These results reinforce the hypothesis presented by Hardie ('87) and Elias and Evans ('83) that histamine functions as a neurotransmitter in the photoreceptors of some, if not all, insect species.