Upregulated expression of oncomodulin, the beta isoform of parvalbumin, in perikarya and axons in the diencephalon of parvalbumin knockout mice.

The calcium-binding proteins parvalbumin, calbindin D-28k, calretinin and calcineurin are present in subsets of GABAergic gigantic calyciform presynaptic terminals of the reticular thalamic nucleus (RTN). Previously it was hypothesized that GABA and calcium-binding proteins including parvalbumin are not only colocalized in the same neuron subpopulation, but that GABA synthesis and parvalbumin expression could be also genetically regulated by a common mechanism. Moreover, parvalbumin expression levels could influence GABA synthesis. For this, we analyzed GABA immunoreactivity in RTN gigantic calyciform presynaptic terminals of parvalbumin-deficient (PV-/-) mice. With respect to GABA immunoreactivity we found no differences compared to wild-type animals. However, using a polyclonal parvalbumin antibody raised against full-length rat muscle parvalbumin on brain sections of PV-/- mice, we observed paradoxical parvalbumin immunoreactivity in partly varicose axons in the diencephalon, mainly in the lamina medullaris externa surrounding the thalamus. A detailed immunohistochemical, biochemical and molecular biological analysis revealed this immunoreactivity to be the result of an upregulation of oncomodulin (OM), the mammalian beta isoform of parvalbumin in PV-/- mice. In addition, OM was present in a sparse subpopulation of neurons in the thalamus and in the dentate gyrus. OM expression has not been observed before in neurons of the mammalian brain; its expression was restricted to outer hair cells in the organ of Corti. Our results indicate that the absence of parvalbumin has no major effect on the GABA-synthesizing system in RTN presynaptic terminals excluding a direct effect of parvalbumin on this regulation. However, a likely homeostatic mechanism is induced resulting in the upregulation of OM in selected axons and neuronal perikarya. Our results warrant further detailed investigations on the putative role of OM in the brain.

The kynurenate analog SZR-72 prevents the nitroglycerol-induced increase of c-fos immunoreactivity in the rat caudal trigeminal nucleus: comparative studies of the effects of SZR-72 and kynurenic acid.

Administration of nitroglycerol in a migraine model results in an increased number of c-fos-expressing secondary sensory neurons in the caudal trigeminal nucleus. Since synapses between first- and second-order trigeminal neurons are mediated by excitatory amino acids, NMDA receptors are inhibited by kynurenic acid, though this crosses the blood-brain barrier only poorly. Systemic treatment of rats with SZR-72, a newly synthetized kynurenic acid analog, diminished the nitroglycerol-induced increase of c-fos immunoreactivity in the brain stem highly significantly, while treatment with kynurenic acid resulted in a significantly smaller decrease, proving that SZR-72 is much more effective than kynurenic acid.

Kynurenine in combination with probenecid mitigates the stimulation-induced increase of c-fos immunoreactivity of the rat caudal trigeminal nucleus in an experimental migraine model.

Nitroglycerin, often used as a migraine model, results in increased number of c-fos immunoreactive secondary sensory neurons in the caudal trigeminal nucleus. Since synapses between first- and second-order trigeminal neurons are mediated by excitatory amino acids, NMDA receptors are presumably inhibited by kynurenic acid, the only known endogeneous NMDA receptor antagonist. Although kynurenic acid does not cross the BBB, its precursor, kynurenine, if combined with probenecid, crosses it readily. Systemic kynurenine + probenecid treatment significantly diminishes nitroglycerin-induced increase of c-fos immunoreactivity in the brainstem.

Decreased expression of kynurenine aminotransferase-I (KAT-I) in the substantia nigra of mice after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment.

Nerve cells in the substantia nigra pars compacta (SNPC) are known to express tyrosine hydroxylase (TH). By means of light and electron microscopical immunohistochemical techniques, we have shown that the dopaminergic neurons of SNPC express also kynurenine aminotransferase (KAT-I), the enzyme taking part in the formation of kynurenic acid, a neuroprotectant which is one of the endogeneous antagonists of N-methyl-d-aspartate receptors. It was also found that microglial cells and astrocytes express KAT-I. It has been shown that the highly selective dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), widely used as a model of Parkinson's disease (PD), affects not only TH of dopaminergic neurons in the SNPC but also their KAT-I immunoreactivity as well: MPTP treatment decreased the number and optical density of KAT-I immunoreactive SNPC neurons. Decrease of KAT-I after MPTP treatment has been proved also by Western blot analysis. MPTP also reduced KAT-I immunoreactivity of microglial cells, except for those involved in reactive gliosis, which were arranged in groups surrounding affected neurons of the SNPC; also the number of KAT-I immunoreactive (IR) astroglial cells was increased in SNPC. We conclude that MPTP treatment may have a dual effect: in addition to being deleterious for neurons expressing TH and KAT-I, it also affects glial cells which could exacerbate the neurodegenerative process characterizing PD.

Glyceryl trinitrate treatment up-regulates soluble guanylyl cyclase in rat dura mater.

Nitric oxide (NO) is a key molecule in vascular headaches and the dura mater has been implicated as a tissue where vascular headache develops. Here we demonstrate expression, enzyme activity and cellular distribution of the intracellular receptor for NO, soluble guanylyl cyclase (sGC), in rat dura mater. Subcutaneous treatment of rats with the NO-donor glyceryl trinitrate (GTN) induced an increase of sGC expression and activity in dural blood vessels after 20-30 min. It has previously been shown that GTN induces headache in normal subjects after 20-30 min. Our findings suggest that an up-regulation of the NO target enzyme contributes to the pathogenesis of GTN-induced headache explaining the subacute rather than acute onset of symptoms.

Functional immunohistochemistry of neuropeptides and nitric oxide synthase in the nerve fibers of the supratentorial dura mater in an experimental migraine model.

The supratentorial cerebral dura of the albino rat is equipped with a rich sensory innervation both in the connective tissue and around blood vessels, which includes nociceptive axons and their terminals; these display intense calcitonin gene-related peptide (CGRP) immunoreactivity. Stereotactic electrical stimulation of the trigeminal (Gasserian) ganglion, regarded as an experimental migraine model, caused marked increase and disintegration of club-like perivascular CGRP-immunopositive nerve endings in the dura mater and induced an apparent increase in the lengths of CGRP-immunoreactive axons. Intravenous administration of sumatriptan or eletriptan, prior to electrical stimulation, prevented disintegration of perivascular terminals and induced accumulation of CGRP in terminal and preterminal portions of peripheral sensory axons. Consequently, immunopositive terminals and varicosities increased in size; accumulation of axoplasmic organelles resulted in the "hollow" appearence of numerous varicosities. Since triptans exert their anti-migraine effect by virtue of agonist action on 5-HT(1D/B) receptors, we suggest that these drugs prevent the release of CGRP from perivascular nerve terminals in the dura mater by an action at 5-HT(1D/B) receptors. Nitroglycerine (NitroPOHL), given subcutaneously to rats, induces increased beading of nitric oxide synthase (NOS)-immunoreactive nerve fibers in the supratentorial cerebral dura mater, and an apparent increase in the number of NOS-immunoreactive nerve fibers in the dural areas supplied by the anterior and middle meningeal arteries, and the sinus sagittalis superior. Structural alterations of nitroxidergic axons innervating blood vessels of the dura mater support the idea that nitric oxide (NO) is involved in the induction of headache, a well-known side effect of coronary dilator agents.

Effects of eletriptan on the peptidergic innervation of the cerebral dura mater and trigeminal ganglion, and on the expression of c-fos and c-jun in the trigeminal complex of the rat in an experimental migraine model.

Nociceptive axons and terminals in the supratentorial cerebral dura mater display an intense calcitonin gene-related peptide (CGRP) immunoreactivity. In an experimental migraine model, it has been shown that electrical stimulation of the rat trigeminal ganglion induced an increase in the lengths of CGRP-immunoreactive axons, increased size and number of pleomorphic axonal varicosities in the dura mater, and an increased number of c-jun and c-fos protein-expressing nerve cells in the trigeminal complex. We demonstrate the effect of the highly specific and moderately lipophilic serotonin agonist eletriptan (Pfizer) which prevents the effects of electrical stimulation in the dura mater. Eletriptan also affected the caudal trigeminal complex; it markedly reduced the numbers of the oncoprotein-expressing cells, mainly after stimulation and to some extent also in nonstimulated animals. Eletriptan also affected expression of CGRP in perikarya of trigeminal ganglion cells, insofar as the number of small nerve cells exhibiting a compact CGRP immunoreaction was decreased to one quarter of the original value. In all these respects, eletriptan acted in a similar way to sumatriptan, with the notable exception that eletriptan also blocked the stimulation-induced effects in the nucleus caudalis trigemini and the upper cervical spinal cord (trigeminal complex), whereas sumatriptan did not. It is concluded that eletriptan, acting on perikarya and both the peripheral and the central axon terminals of primary sensory neurons, exerts its antimigraine effect by an agonist action on 5-HT1B/1D receptors throughout the entire trigeminal system, probably by passing the blood-brain-barrier because of its lipophilic character.

CGRP and adrenomedullin receptor populations in human cerebral arteries: in vitro pharmacological and molecular investigations in different artery sizes.

The aim of the present study was to determine functional and molecular characteristics of receptors for calcitonin gene-related peptide (CGRP) and adrenomedullin in three different diameter groups of lenticulostriate arteries. Furthermore, the presence of perivascular neuronal sources of CGRP was evaluated in these arteries. In the functional studies, in vitro pharmacological experiments demonstrated that both CGRP and adrenomedullin induce alpha-CGRP-(8-37) sensitive vasodilation in artery segments of various diameters. The maximal amounts of vasodilation induced by CGRP and adrenomedullin were not different, whereas the potency of CGRP exceeded that of adrenomedullin by 2 orders of magnitude. Significant negative correlations between artery diameters and maximal responses were demonstrated for CGRP and adrenomedullin. In addition, the potency of both peptides tended to increase in decreasing artery diameter. In the molecular experiments, levels of mRNAs encoding CGRP receptors and receptor subunits were compared using reverse transcriptase polymerase chain reactions (RT-PCR). The larger the artery, the more mRNA encoding receptor activity-modifying proteins 1 and 2 (RAMP1 and RAMP2) was detected relative to the amount of mRNA encoding the calcitonin receptor-like receptor. By immunohistochemistry, perivascular CGRP containing nerve fibres were demonstrated in all the investigated artery sizes. In conclusion, both CGRP and adrenomedullin induced vasodilation via CGRP receptors in human lenticulostriate artery of various diameter. The artery responsiveness to the CGRP receptor agonists increased with smaller artery diameter, whereas the receptor-phenotype determining mRNA ratios tended to decrease. No evidence for CGRP and adrenomedullin receptor heterogeneity was present in lenticulostriate arteries of different diameters.

Effect of the pesticide Deltamethrin on the Mauthner cells of Lake Balaton fish.

Deltamethrin, a synthetic pesticide [(S)alpha-cyano-3-phenoxybenzyl-(1R)-cis-3-(2.2-dibromovinyl)-2,2-dim ethylcyclopropane-carboxylate] used for extermination of mosquitoes on the shores of lake Balaton, has been found to induce severe impairments of the nervous system of several Lake Balaton fish, such as carp (Cyprinus carpio), goldfish (Carassius auratus gibelis Bloch), eel (Anguilla anguilla) and wels (Silurus glanis). It has been shown that Deltamethrin, in a concentration of 1 microgram/liter in the aquarium water, inhibits acetylcholinesterase enzyme activity of the giant Mauthner's nerve cells as well as of the axon terminals synapsing with these cells. Even more importantly, however, Deltamethrin in a concentration of 10 micrograms per liter, induces blockade of the expression of choline acetyltransferase in the bulbous axon terminals synapsing with the lateral dendrites of the Mauthner cells. Since, under normal conditions, the function of the Mauthner cells is to co-ordinate the C-start reaction, by which fish rapidly leave sites of nociceptive stimulation, it stands for reason to assume that Deltamethrin intoxicated fish may be prone to become victims of various factors which endanger survival of the individual. During the last decade, waves of fish deaths were observed in Lake Balaton, which is the largest fresh-water lake in Europe. Fish death coincided with airborne mosquito-killing campaigns. Results of the enzyme- and immunohistochemical studies described in this paper, together with the deleterious effects of Deltamethrin to the enteric nervous system of fish which has been reported earlier (Lang et al., 1997) suggest that fish death might be caused by the indiscriminate use of Deltamethrin airborne spray in the mosquito-extermination campaigns.

Effects of in vivo sodium azide administration on the immunohistochemical localization of kynurenine aminotransferase in the rat brain.

Endogenous excitotoxins that act on receptors of cerebral excitatory amino acids play important roles in the pathogenesis of excitotoxic brain diseases. Activation of excitatory amino acid receptors results in neuronal death characteristic of these disorders. Kynurenic acid, a powerful endogenous excitatory amino acid receptor antagonist, which is therefore widely regarded as a potent neuroprotective agent, is produced from its biological precursor, L-kynurenine, by the action of the enzyme kynurenine aminotransferase-I. The chemical hypoxia induced by mitochondrial toxins produces a secondary excitotoxicity, leading to the activation of N-methyl-D-aspartate receptors. Accordingly, sodium azide, an inhibitor of cytochrome oxidase, induces the release of excitotoxins via an energy impairment and this, in turn, results in neurodegeneration. Since energy-dependent secondary excitotoxic mechanisms also account for the pathogenesis of neurodegenerative diseases, a study was made of the effects of sodium azide on the immunohistochemical localization of kynurenine aminotransferase-I. After in vivo administration of sodium azide for five days, a markedly decreased glial kynurenine aminotransferase-I immunoreactivity was found by immunohistochemical techniques in the glial cells of the striatum, hippocampus, dentate gyrus and temporal cortex; at the same time, kynurenine aminotransferase-I started to be expressed by nerve cells which had not been immunoreactive previously. The accumulation of kynurenine aminotransferase-I reaction product around the ribosomes of neuronal endoplasmic reticulum suggests de novo synthesis of kynurenine aminotransferase-I in the reactive nerve cells.

Parabrachial origin of calcitonin gene-related peptide-immunoreactive axons innervating Meynert's basal nucleus.

Meynert's basal nucleus is innervated by calcitonin gene-related peptide (CGRP)-immunoreactive axons synapsing with cholinergic principal cells. Origin of CGRP-immunopositive axons was studied in the albino rat. Since beaded axons containing the nicotinic acetylcholine receptor (nAChR) are also present in the basal nucleus, the microstructural arrangement raises the question whether or not an interaction between CGRP and nAChR exists like in the neuromuscular junction. We found that electrolytic lesion of the parabrachial nucleus results in degeneration of CGRP-immunoreactive axons in the ipsilateral nucleus basalis and induces shrinkage of principal cholinergic neurons while the contralateral nucleus basalis remains intact. Electrolytic lesions in the thalamus, caudate-putamen, and hippocampus did not induce alterations in Meynert's basal nucleus. Disappearance of CGRP after lesions of the parabrachial nucleus does not impair presynaptic nAChR in the basal nucleus, suggesting that, unlike in the neuromuscular junction, CGRP is not involved in the maintenance of nAChR in the basal forebrain. It is concluded that the parabrachial nucleus is involved in the activation of the nucleus basalis-prefrontal cortex system, essential in gnostic and mnemonic functions.

Infraterminal spreading and extrajunctional expression of nicotinic acetylcholine receptors in denervated rat skeletal muscle.

Through the use of biotinylated-bungarotoxin and monoclonal antibodies, the nicotinic acetylcholine receptor (nAChR) was localized in the subneural apparatus of mammalian motor end plates of the flexor digitorum brevis muscle of the adult rat at the light and electron microscopic levels. Under normal conditions, nAChR was located in the primary post-synaptic membrane of the neuromuscular junction, and the depths of the junctional folds constituting the secondary post-synaptic membrane did not contain any nAChR. Up to 75 days after repeated transection of the related motor nerve (sciatic), there was no major alteration in the light-microscopic localization of junctional nAChR in the subneural apparatus, except for a moderate shrinkage and increased immunocytochemical reactivity of the subneural apparatus. At the electron microscopic level, however, immunocytochemical reactivity gradually occupied the entire extent of the secondary post-synaptic membrane, including the depths of the junctional folds, which exhibited extensive branching. In non-innervated portions of the muscle fibers, nAChR receptor appeared in a linear localization on the surfaces of denervated muscle fibers. This linear reaction was not continuous with the nAChR reaction of the motor end plates. It is concluded that denervation supersensitivity might not be due to spreading of junctional nAChR from the end-plate area, but rather to expression of nAChR in non-innervated portions of the muscle fiber and to the infraterminal (subsynaptic) spreading of nAChR into the depths of junctional folds.

Illusive transience of parvalbumin expression during embryonic development of the primate spinal cord.

Parvalbumin has been located by pre-embedding light- and electron microscopic immunohistochemical techniques in the spinal cords of monkey fetuses (Macaca fasciculata), ranging from E70 to E 123, and in young (P20) and young adult (3 years) Macaque monkeys. During the time window investigated, the main developmental events of parvalbumin-containing neural elements are that parvalbumin-positive dorsal root collaterals establish intercellular networks first around nerve cells of Clarke's nucleus, then in the motoneuron pool and finally in the upper dorsal horn. In each of these areas, location of the parvalbumin-positive network is gradually shifted from medial to lateral. Whenever an intercellular network is established, nerve cells innervated by parvalbumin-positive terminals of dorsal root collaterals start to express parvalbumin. Immunoreactivity of dorsal root axons is transient; it disappears first from the intercellular networks and, afterwards, also from the dorsal columns. However, the pericellular synaptic terminals and their post-synaptic nerve cells express parvalbumin into adulthood. It is concluded that some of the large (Type A) dorsal root ganglion cells are the first ones in the spinal reflex pathway to express parvalbumin, which is elicited and gradually increased in nerve cells synaptically innervated by parvalbumin-positive axon terminals. This seems to represent a specific case of activation (or desinhibiton) of the genome. Apparent "transience" of parvalbumin is due to the specific geometry of primary sensory neurons equipped with extremely long axonal processes, and the consequent specialities of axonal transport characteristics.

Effect of a nitric oxide donor on nitroxergic nerve fibers in the rat dura mater.

Nitroglycerine, given subcutaneously to rats (10 mg/kg body weight) induces increased beading of nitric oxide synthase immunoreactive (NOS-IR) nerve fibers in the supratentorial cerebral dura mater, and an apparent increase in the number of NOS-IR nerve fibers in the dural areas supplied by the anterior and middle meningeal arteries, and the sinus sagittalis superior. Structural alterations of nitroxergic axons innervating blood vessels of the dura mater support the idea that nitric oxide is involved in the induction of headache also by a primary peripheral action, a well-known side effect of coronary dilator agents.

Development of glomerular synaptic complexes and immunohistochemical differentiation in the superficial dorsal horn of the embryonic primate spinal cord.

Development of glomerular synapses in the superficial dorsal horn has been studied in the embryonic macaque spinal cord using light and electron microscopic techniques including Golgi impregnation, 3H-thymidine radioautography and pre-embedding immunohistochemistry of substance P (SP), calcitonin gene related peptide (CGRP), calbindin D-28 K (CB) and parvalbumin (PV). The study revealed that substantia gelatinosa cells of the primate dorsal horn are generated last, but unlike in rodents, synaptogenesis in this region starts at early embryonic (E) stages of the 165-day long gestation. Already by E30, both Type 1 (light) and 2 (dark) dorsal root axons and their growth cones are identifiable within the oval bundle of His, before they form synaptic contact with their final target cells. Subsequently they invade the dorsal horn and enter the bisecting interfaces formed by orderly programmed cell death. Each type of scalloped (sinusoid) central primary afferent terminal (i.e. DSA, RSV and LDCV) have well defined pre- and post-synaptic specializations already by E40. Among the neuropeptides studied, SP appears first at E67 and CGRP at E70 in the lateral position but within a few days both of them are spread to the entire superficial dorsal horn. Both SP and CGRP are present in the thin dorsal root axons and their growth cones, giving rise to scalloped and simple axon terminals. PV is transiently present in the entire length of the thick dorsal root afferents before becoming concentrated in the synaptic boutons. CB is displayed mainly in neurons of the lamina I and III. Dendrites of CB-immunoreactive cells establish synaptic connection with each type of dorsal root afferents, including glomerular synaptic complexes. These data reveal that the superficial dorsal horn in the primate spinal cord develops its characteristic synaptic complexes much earlier in gestation than in any other mammalian species studied. Furthermore, characteristic cytological features of the prospective glomerular complex emerge before establishment of the final synaptic contacts.

Peptidergic innervation and the nicotinic acetylcholine receptor in the primate basal nucleus.

Peptidergic innervation and localization of the neuronal nicotinic acetylcholine receptor (nAChR) was studied in the basal forebrain of Macaca fascicularis in order to provide microstructural proofs for the theory (Changeux et al., 1992) that calcitonin gene-related peptide (CGRP) is responsible for the maintenance of the acetylcholine receptor. Distribution and localization of five neuropeptides, namely substance P (SP), CGRP, neuropeptide Y (NPY), vasoactive intestinal polypeptide (VIP) neurotensin (NT), and the neuropeptides parvalbumin (PV) and the alpha-bungarotoxin- (alpha-BTX-) binding protein was studied by means of light- and electron microscopic pre-embedding immunocytochemistry. Immunohistochemical double staining revealed that large cholinergic principal nerve cells in the basal forebrain, corresponding to cell group Ch4 constituting Meynert's basal nucleus (BNM), and exerting intense choline acetyltransferase (ChAT) immunoreactivity, are synaptically innervated by axons displaying CGRP immunoreactivity. While SP, NPY, PV and CGRP establish dense networks in BNM, innervation by NT and VIP is sparse. Biotinylated alpha-BTX visualizes beaded axons that surround dendrites and perikarya of cholinergic principal cells. Electron microscopic organization of the neuropil in BNM is characterized by a glomerular (or rather cartridge-like) arrangement of axons surrounding dendrites of non-cholinergic principal nerve cells. At least one of the axons establishing the glomerulus (cartridge) exerts CGRP immunopositivity while alpha-BTX-immunopositive axons, presynaptic to dendrites of principal cells, are attached to the glomeruli (cartridges) from outside. As alpha-BTX-binding indicates localization of the alpha7 subunit of the neuronal nAChR, the microtopographical arrangement supports the idea that, in a manner similar to that in the neuromuscular junction, CGRP might contribute to the maintenance of nAChR also in BNM. Our results suggest that presynaptic nAChR-s are involved in the regulation of acetylcholine release from a feed-forward amplification mechanism of cholinergic principal cells of BNM.

Depletion of calcitonin gene-related peptide from the caudal trigeminal nucleus of the rat after electrical stimulation of the Gasserian ganglion.

Electrical stimulation of the Gasserian ganglion resulted in partial depletion of calcitonin gene-related peptide (CGRP) from ipsilateral central terminals of pseudounipolar primary sensory ganglion cells. Affected terminals exhibit decreased CGRP immunoreactivity as shown by cytophotometric densitometry of the caudal trigeminal nucleus. The decrease in CGRP immunoreactivity is statistically significant only in the medial one-third of the caudal trigeminal nucleus. Since earlier studies have shown that electrical stimulation of the Gasserian ganglion induces first accumulation then depletion of CGRP from perivascular sensory terminals in the dura mater, the present experiments suggest that CGRP is depleted also from central terminals of primary sensory trigeminal neurons, which might be of importance in the pathogenesis of migraine headache.

Nitric oxide synthase and the acetylcholine receptor in the prefrontal cortex: metasynaptic organization of the brain.

Nitric oxide synthase (NOS) and the nicotinic acetylcholine receptor (nAChR) immunoreactivity of the cerebral cortex was studied in adult Macaca fascicularis monkeys at light- and electron microscopic levels. NOS was located by means of the polyclonal antibodies developed by Transduction Laboratories (Lexington, KY, USA), as primary serum, in a dilution of 1:1000, and nAChR was located by means of biotinylated alpha-bungarotoxin (BTX) obtained from Molecular probes (Eugene, Oregon, USA) in a dilution of 1:2000. While endothelial eNOS outlined blood vessels in the brain, brain-derived (neural) bNOS labelled three well-defined cell types in area 46 of the prefrontal cortex, viz. (a) bipolar cells, scattered through layers III to V, equipped with long dendrites which pass over the thickness of the cortex in a right angle to the pial surface, establishing dendritic bundles closely reminiscent of a columnar organization; (b) large multipolar cells, located mainly in layers V and VI, with axons which interconnect dendritic bundles of the bipolar cells and establish synapses with dendritic shafts and spines of the former; and (c) stellate cells, located in lamina II and III, which establish an axonal network in lamina zonalis (lamina I). This arrangement is most characteristic in area 46 of the prefrontal cortex; areas 10 and 12 display similar features. In contrast, the primary visual cortex (area 17), is lacking any sign of columnar organization. Localization of bNOS immunoreactivity is at marked variance to that of NADPH-diaphorase which labels large pyramidal cells in the primate cortex. Binding of alpha-bungarotoxin (BTX) which labels the alpha 7 subunit of nAChR is located in somata, dendrites and axons of interneurons scattered over the entire width of the prefrontal cortex; on the other hand, the monoclonal antibody mAb 35 which labels subunits alpha 1, alpha 3 and alpha 5 in the main immunogenic region of the receptor, visualizes apical dendritic shafts similar to those like bNOS. Strategic localization of bNOS in the primate prefrontal cortex fulfills criteria of producing a freely diffusing retrograde messenger molecule operative in signal transduction routes subserving topography and columnar organization of the cortex, as well as long-term potentiation and long-term depression phenomena underlying mnemonic and gnostic functions. Common occurrence of bNOS and nAChR in identical or similar structures in the prefrontal cortex suggests that interactions between nitrogen oxide and presynaptically released acetylcholine might be involved in the metasynaptic organization of the cerebral cortex, operating in a non-synaptic manner in maintaining optimal performance on cognitive tasks.

Effect of a serotonin agonist (sumatriptan) on the peptidergic innervation of the rat cerebral dura mater and on the expression of c-fos in the caudal trigeminal nucleus in an experimental migraine model.

The supratentorial cerebral dura of the albino rat is equipped with a rich sensory innervation including nociceptive axons and their terminals, which display intense calcitonin gene-related peptide (CGRP) immunoreactivity both in the connective tissue and around blood vessels. Stereotactic electrical stimulation of the trigeminal (Gasserian) ganglion, regarded as an experimental migraine model, induces marked increase and disintegration of club-like perivascular CGRP-immunopositive nerve endings in the dura. Intravenous administration of sumatriptan, prior to electrical stimulation, prevents disintegration of perivascular terminals and induces accumulation of CGRP in terminal and preterminal portions of peripheral sensory axons. Consequently, immunopositive terminals and varicosities increase in size; accumulation of axoplasmic organelles results in a "hollow" appearance of many varicosities. Since sumatriptan exerts its anti-migraine effect by virtue of its agonist action on 5-HT1D receptors, we suggest that sumatriptan prevents the release of CGRP from dural perivascular terminals by an action at 5-HT1D receptors. In the caudal trigeminal nucleus electrical stimulation of the trigeminal ganglion induces, in interneurons, increased expression of the oncoprotein c-fos which is not prevented by intravenous application of sumatriptan. Disparate findings regarding this effect are partly due to the fact that sumatriptan very poorly passes the blood-brain barrier and partly to different experimental paradigms used by different authors.