Tetrahydrobiopterin modulates the behavioral neuroinflammatory response to an LPS challenge in mice.

Tetrahydrobiopterin (BH4) is a necessary cofactor for the synthesis of monoamines from essential amino-acids, phenylalanine, tyrosine and tryptophan. The BH4 synthesis pathway is induced by inflammatory factors but highly regulated processes maintain levels in a physiological range. However, BH4 activity can be durably altered in inflammation-related pathologies, such as certain types of depression, potentially involving impairment of dopaminergic neurotransmission. The purpose of this study was to investigate the response of the brain BH4 pathway to the inflammatory stimulus induced by lipopolysaccharide (LPS) in mice. Brain expression of genes related to BH4 synthesis, levels of BH4, changes in L-aromatic amino acid precursors of monoamines and dopamine levels were determined. As secondary aim, the effect of acute BH4 supply under the inflammatory challenge was tested on these parameters and on the expression of inflammatory cytokines. Mice were also submitted to the sucrose preference test and to the open-field in order to asses hedonic and locomotor responses to LPS, in addition to their modulation by BH4 supply. The LPS challenge resulted in decreased striatal DA levels and increased Phenylalanine/Tyrosine ratio, suggesting reduced BH4 activity. BH4 supply was effective to increase striatal BH4 levels, to restore the LPS-induced decreased in DA levels in striatum and to dampen the LPS-induced expression of inflammatory cytokines. At the behavioral level, BH4 supply was able to restore the loss of locomotor response to amphetamine in the LPS treated mice, suggesting a modulation of the dopaminergic neurotransmission. These data suggest that BH4 can be considered as a potential add-on molecule, helping to maintain or restore dopaminergic neurotransmission in neuroinflammatory conditions..

Tetrahydrobioterin (BH4) Pathway: From Metabolism to Neuropsychiatry.

Tetrahydrobipterin (BH4) is a pivotal enzymatic cofactor required for the synthesis of serotonin, dopamine and nitric oxide. BH4 is essential for numerous physiological processes at periphery and central levels, such as vascularization, inflammation, glucose homeostasis, regulation of oxidative stress and neurotransmission. BH4 de novo synthesis involves the sequential activation of three enzymes, the major controlling point being GTP cyclohydrolase I (GCH1). Complementary salvage and recycling pathways ensure that BH4 levels are tightly kept within a physiological range in the body. Even if the way of transport of BH4 and its ability to enter the brain after peripheral administration is still controversial, data showed increased levels in the brain after BH4 treatment. Available evidence shows that GCH1 expression and BH4 synthesis are stimulated by immunological factors, notably pro-inflammatory cytokines. Once produced, BH4 can act as an anti- inflammatory molecule and scavenger of free radicals protecting against oxidative stress. At the same time, BH4 is prone to autoxidation, leading to the release of superoxide radicals contributing to inflammatory processes, and to the production of BH2, an inactive form of BH4, reducing its bioavailability. Alterations in BH4 levels have been documented in many pathological situations, including Alzheimer's disease, Parkinson's disease and depression, in which increased oxidative stress, inflammation and alterations in monoaminergic function are described. This review aims at providing an update of the knowledge about metabolism and the role of BH4 in brain function, from preclinical to clinical studies, addressing some therapeutic implications.

Lipopolysaccharide-induced depressive-like behavior is mediated by indoleamine 2,3-dioxygenase activation in mice.

Although elevated activity of the tryptophan-degrading enzyme indoleamine 2,3-dioxygenase (IDO) has been proposed to mediate comorbid depression in inflammatory disorders, its causative role has never been tested. We report that peripheral administration of lipopolysaccharide (LPS) activates IDO and culminates in a distinct depressive-like behavioral syndrome, measured by increased duration of immobility in both the forced-swim and tail suspension tests. Blockade of IDO activation either indirectly with the anti-inflammatory tetracycline derivative minocycline, that attenuates LPS-induced expression of proinflammatory cytokines, or directly with the IDO antagonist 1-methyltryptophan (1-MT), prevents development of depressive-like behavior. Both minocycline and 1-MT normalize the kynurenine/tryptophan ratio in the plasma and brain of LPS-treated mice without changing the LPS-induced increase in turnover of brain serotonin. Administration of L-kynurenine, a metabolite of tryptophan that is generated by IDO, to naive mice dose dependently induces depressive-like behavior. These results implicate IDO as a critical molecular mediator of inflammation-induced depressive-like behavior, probably through the catabolism of tryptophan along the kynurenine pathway.

Different responsiveness of spleen lymphocytes from two lines of psychogenetically selected rats (Roman high and low avoidance).

Roman high- (RHA) and low-avoidance (RLA) rats have been genetically selected on the basis of their active avoidance behavior, and have been shown to differ on numerous behavioral, neurochemical and neuroendocrine parameters, especially in response to stress. We investigated the activity of splenic lymphocytes in vitro. Natural killer cell activity against YAC-1 tumoral cells and the mitotic response to plant lectins concanavalin A and phytohemagglutinin were much lower for lymphocytes isolated from RHA rats, in males as well as in females. The difference between the two strains was even larger when measured in a stressed state, immediately after active avoidance learning. On the other hand, the mitotic response to bacterial lipopolysaccharide, a B-cell-specific mitogen, was not different between the two lines, indicating that the difference in lymphocyte reactivity is limited to the T-lineage. The lower activity of T-cells in the RHA line had no consequence upon the ability of these animals to build up an antibody response against sheep red blood cells. These results indicate that Roman lines are an interesting animal model for the study of the relationships between the brain and the immune system, as well as for the analysis of the genes involved in the control of behavior.

Psychobiogenetics: adapted tools for the study of the coupling between behavioral and neuroendocrine traits of emotional reactivity.

The adaptive response to environmental challenges involves both behavioral and neuroendocrine adjustments. Several examples show that the behavioral and neuroendocrine responses to environmental stimulations, far from being independent, are frequently interrelated and/or coselected. The existence of such a coselection in genetically selected models provides an advantageous tool for the study of genetic influence on emotional reactivity and the mechanism of linkage between behavioral and neuroendocrine traits. In particular, the study of the Roman rat lines which have been selected for their performance in active avoidance behavior has allowed us to establish the respective role of genetic and maturation factors in the reactivity of these animals to stress. Indeed, the role of maturation in determining interindividual differences appears to be more marked in RLA than in RHA rats. The study of genetic factors, which play a major role in shaping individual characteristics of reactivity, may help, therefore, to elucidate the biological mechanisms coupling behavioral and neuroendocrine aspects of personality structure to emotional reactivity.

Central injection of IL-10 antagonizes the behavioural effects of lipopolysaccharide in rats.

Peripheral (i.p.) and central (i.c.v.) injections of lipopolysaccharide (LPS) have been shown to induce brain expression of proinflammatory cytokines and to depress social behaviour in rats, increase duration of immobility and induce body weight loss. To determine if the anti-inflammatory cytokine, interleukin-10 (IL-10) is able to modulate these effects, recombinant rat IL-10 was injected in the lateral ventricle of the brain (30, 100, 300 ng/rat) prior to i.p. or i.c.v. injection of LPS (250 micrograms/kg or 60 ng/rat, respectively). Social exploration was depressed for 6 h after i.p. LPS injection. This effect was attenuated by IL-10 (30 and 100 ng) 2 h after injection, whereas the highest dose of IL-10 blocked the depression of social interaction for 6 h after LPS injection. IL-10 produced the same effects on the increase of immobility although the results did not reach significance. Social exploration was depressed 3 h after i.c.v. LPS injection, and this was accompanied by increased immobility. These effects were totally blocked by i.c.v. IL-10 (300 ng/rat). Rats lost body weight after i.c.v. LPS, and this effect was attenuated by i.c.v. IL-10. These results indicate that IL-10 is able to modulate the production and/or action of central proinflammatory cytokines.

Chronic treatment with the atypical antidepressant tianeptine attenuates sickness behavior induced by peripheral but not central lipopolysaccharide and interleukin-1beta in the rat.

RATIONALE: The hypothesis that proinflammatory cytokines play a causative role in the pathophysiology of depression has been recently tested by studying the effect of antidepressants on production of endogenous cytokines, and on sickness behavior induced by exogenous cytokines. In this last case, however, the effect of antidepressants has been only studied on the effect of peripherally administered cytokines. OBJECTIVES: The aim of the present study was to determine whether the antidepressant tianeptine can attenuate both peripheral and central cytokine actions. METHODS: Rats were injected IP with acute (10 mg/kg) or chronic (10 mg/kg, 2 times/day, 17 days) tianeptine. The effects of this treatment were assessed on the behavioral (social exploration, locomotion) and metabolic (food intake, body weight) alterations induced by peripheral or central administration of the cytokine inducer lipopolysaccharide (LPS) (250 microg/kg IP; 100 ng/rat ICV) or the prototypical proinflammatory cytokine interleukin-1 (IL-1)beta (15 microg/rat IP; 90 ng/rat ICV). RESULTS: Chronic, but not acute, treatment with tianeptine attenuated the behavioral signs of sickness behavior induced by peripheral, but not central, LPS or IL-1beta. CONCLUSIONS: This work, which is the first in vivo study assessing the effect of an antidepressant on centrally induced immune activation, shows a clear dissociation between peripheral and central cytokine effects, and suggests a peripheral site of action of tianeptine. It also provides the first evidence that the protective effects of classical antidepressants on LPS-induced sickness behavior extend to an atypical antidepressant, and that the protective effect of antidepressants also applies to IL-1beta.

Cerebral tryptophan hydroxylase activity, and 5-HT1A receptor, 5-HT2A receptor, and 5-HT transporter binding in grouped and isolated Roman RHA and RLA rats: relationships with behaviours in two models of anxiety.

Male Roman low-(RLA) and high-avoidance (RHA) rats differ when tested in the elevated plus-maze and the black/white box, but not when (isolated and) tested for their social interaction. Herein, we have analysed the impact of prior isolation on male Roman rats tested in the first two models of anxiety; moreover, because central serotonin (5-HT) systems in Roman rats have been scarcely studied, we have also analysed several anxiety-related indices of central serotonergic activity in grouped/isolated Roman rats. Group-housed RLA rats tested in the elevated plus-maze and the black/white box were less anxious than their RHA counterparts, thereby confirming our previous study. Isolation had anxiogenic (and hypolocomotor) effects, these being significant in RLA rats only. Tryptophan hydroxylase activity in midbrain (but not in cortex, hippocampus or hypothalamus) was lower in group-housed (but not in isolated) RLA rats than in RHA rats, a difference independent from changes in the regulatory properties of the enzyme. Neither midbrain and hippocampal [3H]8-hydroxy-2-(di-n-propylamino)-tetrlin binding at 5-HT1A receptors, nor midbrain [3H] citalopram binding at the 5-HT transporter was different between grouped/isolated RHA/RLA rats. Alternatively, a trend toward a lower hypothalamic [3H]citalopram binding in (group-housed) RLA rats than in RHA rats could be noted, whereas cortical [3H]ketanserin binding at 5-HT2A receptors was lower in RLA rats than in RHA rats, a difference prevented by prior isolation. This study opens the possibility that inter-line differences in 5-HT2A receptors partly (or totally) underlie the respective behaviours of RHA and RLA rats in the elevated plus-maze and the black/white box.

Regulation of [3H]5-HT release in raphe, frontal cortex and hippocampus of 5-HT1B knock-out mice.

Regulation of the electrically evoked release of [3H]5-HT was examined in midbrain, frontal cortex and hippocampus preloaded slices obtained from wild-type and 5-HT1B knock-out mice. In the absence of any drug [3H]5-HT released was increased in midbrain and hippocampus but not in frontal cortex slices of [3H]5-HT1B knock out mice. The selective 5-HT1B agonist CP 93129 and the 5-HT1B/1D agonist sumatriptan, inhibited [3H]5-HT release in hippocampus and cortical slices obtained from control mice but had no effect in mutants. In the two projection areas studied, the non-selective 5-HT agonist 5-carboxyamidotryptamine (5-CT) inhibited [3H]5-HT release in both groups of mice, indicating that additional 5-HT receptors, other than 5-HT1B, might be involved in the regulation of [3H]5-HT release from 5-HT terminals. In slices containing midbrain raphe nuclei, CP 93129 had no effect in either group. In contrast, sumatriptan inhibited [3H]5-HT release in controls and mutants. The latter effect was blocked by the 5-HT1D antagonist GR 127935, but not the 5-HT1A antagonist (+)WAY 100135, thus suggesting that a 5-HT1D-like receptor, possibly 5-HT1D alpha, negatively regulates 5-HT release in mouse midbrain raphe nuclei.

Cocaine sensitivity in Roman High and Low Avoidance rats is modulated by sex and gonadal hormone status.

Repeated exposure to stressful stimuli or psychostimulants increases sensitivity to the motoric effects of these drugs, i.e. behavioral sensitization. The objective of the present experiment was to determine the interaction between factors that modulate psychostimulant sensitivity, i.e. sex and circulating gonadal hormone levels, in rats with a genetically distinct locomotor response to novelty: Roman Low Avoidance rats (RLA) freeze while Roman High Avoidance rats (RHA) remain motorically active. Ninety-six male and female RHA and RLA rats were gonadectomized (GDX) just after weaning or as adults, or left gonadally intact. Each rat received a total of 9 injections of cocaine hydrochloride (10 mg/kg, IP), at 3-4 day intervals for 5 weeks. Locomotor activity was measured after each injection, and stereotypes were rated 1 x/week. Open field behavior (10 min) and plasma corticosterone were measured 2 weeks after the final injection. Overall, the RHA line was more sensitive to (1) cocaine's stereotypic effects, and (2) the influence of ovarian hormones on the cocaine's acute and sensitizing effects on locomotor activity. Therefore, genetic background not only determines cocaine sensitivity, but also the influence of gonadal hormones on locomotor activity. These interactions are relevant when considering the genetic contribution to abuse liability.

Male Fischer 344 and Lewis rats display differences in locomotor reactivity, but not in anxiety-related behaviours: relationship with the hippocampal serotonergic system.

Recent studies have shown that arthritis-susceptible Lewis female rats display a marked hypoactivity of the hypothalamo-pituitary-adrenal (HPA) axis and decreased concentrations of hippocampal serotonin receptors (5-HT1A), when compared with arthritis-resistant Fischer 344 female rats. Although previous studies have suggested that these inter-strain differences may extend to several behaviours, the hypothesis that Fischer 344 and Lewis differ in their anxiety and locomotor scores when placed in novel environments has been only scarcely tested. The present study has thus analysed the behaviours of male Fischer 344 and Lewis rats placed successively in activity cages, in an open field (low and high aversive conditions), and in two animal models of anxiety (the elevated plus-maze, the black/white box). Moreover, because the present study was conducted with male rats, we have also checked whether the HPA axis- and 5-HT1A receptor-related differences previously described between female Fischer 344 and Lewis rats extended to males. Under basal conditions: (i) activity of the HPA axis; and (ii) hippocampal 5-HT1A receptor binding and activity of tryptophan hydroxylase (the rate-limiting enzyme in 5-HT biosynthesis) were decreased in Lewis rats, compared with Fischer 344 rats. In addition, the response of the HPA axis to a mild stress (10 min in a novel environment) was lower in Lewis rats than in Fischer 344. When placed in activity cages, Lewis rats displayed a lower locomotor activity, compared with Fischer 344 rats. In the open-field, Lewis rats cross a lower number of inner squares and groomed less than Fischer 344 rats. In the elevated plus-maze and in the black/white box, Fischer 344 and Lewis rats exhibited similar 'anxious' profiles as none of the rats visited the open arms (elevated plus-maze) and the white compartment (black/white box). This study, which extends earlier neurochemical and neuroendocrine findings in females, suggests that both strains display high levels of anxiety but markedly differ in their locomotor activities. Whether the latter strain difference is due to alterations in the HPA axis and/or the central serotonergic systems is an issue that remains to be explored.

Paradoxical differences in animal models of anxiety among the Roman rat lines.

The Roman high-avoidance (RHA) and low-avoidance (RLA) rat lines have been selected upon their divergent active avoidance behaviours. On the basis of open-field behavioural analyses, it has been suggested that RLA rats display more emotionality than RHA rats. Herein, we have analysed the behaviours of male RHA and RLA rats in three tests putatively related to anxiety, namely the elevated plus-maze, the black/white box, and the social interaction test. In the elevated plus-maze, neither the number of total arm entries nor the percent number of open arm entries were different between the Roman lines. Alternatively, RLA rats spent more time on the open arms, compared to RHA rats. In the black/white box, both the latency to enter the black compartment, the number of shuttles, the time spent in the white compartment, and the general activity in the white compartment were higher in RLA rats, compared to their RHA counterparts. Lastly, socially isolated RHA and RLA behaved similarly when exposed to a social interaction test. It is suggested that under particular experimental conditions male RLA rats display less anxiety than male RHA rats, and that the open-field test may provide indices of activity rather than indices of anxiety.

Maturation of the behavioral and neuroendocrine differences between the Roman rat lines.

The behavioral and/or neuroendocrine reactivity to psychological (open-field exposure) and physiological (CRF challenge) stimulations, as well as adrenal tyrosine hydroxylase (TH) and phenylethanolamine N-methyl transferase (PNMT) activities were measured, at different ages, in the Roman high avoidance (RHA) and Roman low avoidance (RLA) rat lines that have been genetically selected on the basis of their divergent active avoidance behavior. The highest locomotor activity in the open field, associated to blunted prolactin and renin reactivity to an emotional stress and lower specific TH and PNMT activities, characterized the RHA rats of all ages. HPA axis reactivity to psychological and/or physiological stimulations was identical in young animals (14 weeks old) of the two lines. Nevertheless, it displayed with age maturation processes, since the amplitude of postopen-field secretion peak for ACTH was larger in RLA rats from 20 weeks on, the response to CRF being not increased until 42 weeks. These maturation processes could result from genetically influenced changes related to environmental stimulations. Therefore, the Roman lines may be an excellent model to study the interactions between the genetic and developmental factors controlling the coupling between both behavioral and neuroendocrine functions.

Prolactin as a link between behavioral and immune differences between the Roman rat lines.

Roman high (RHA)- and low (RLA)-avoidance rats are two lines of Wistar rats genetically selected on the basis of their active avoidance behavior in a shuttle-box. They also differ in several other behavioral responses, such as their locomotor activity in novel environments (open-field, circular corridor), with the RHA rats being more active than the RLA animals, as well as in endocrine reactivity and immune functions. These experiments were designed to investigate further the neuroendocrine characteristics of these animals as a possible link between the brain and immune functions. Despite the marked behavioral and immune differences observed, no between-lines variation could be found in basal hypothalamo-pituitary-adrenocortical axis activity or in its responses to different protocols of novel environment stress, or after corticotropin-releasing factor (CRF) challenge. On the other hand, stimulated prolactin levels were higher in the low avoidance line. These results exclude the pituitary-adrenocortical axis and suggest prolactin as a link between behavioral and immune differences between the Roman lines. Moreover, these results indicate that these rats may be an excellent model for the study of the relationships between the brain and the immune system.

Multiple neuroendocrine responses to chronic social stress: interaction between individual characteristics and situational factors.

After four weeks of individual housing, male Wistar rats (selected for high or low spontaneous aggressiveness by multiple round-robin encounters) were housed three per cage and submitted to four weeks of chronic social stress consisting of changing membership in the social groups by daily rotation of the animals among cages every day according to a random permutation procedure. In addition, half the males in each condition were housed with three females. Each environmental condition triggered different neuroendocrine changes. Cohabitation with females increased the hypothalamo-pituitary-adrenocortical axis activity, including enlargement of adrenals and increased circulating corticosterone levels. On the other hand, daily rotation of the rats between different social groups activated part of the sympathetic nervous system, such as increased phenylethanolamine N-methyl transferase (PNMT) activity in the adrenals. The level of aggressiveness, however, had no direct influence but interacted with environmental factors on such neuroendocrine measures as circulating testosterone or plasma renin activity. These results indicate that during chronic stress, there is no single, unique response by the animal, but a highly complex set of neuroendocrine changes, dependent on the interaction between individual characteristics (the level of aggressiveness is an example) and situational factors.

Modulation of the effects of cocaine by 5-HT1B receptors: a comparison of knockouts and antagonists.

Serotonergic transmission has been suggested to modulate the effects of cocaine. However, the specific receptors underlying this phenomenon have not been identified. To evaluate the role of the 5-HT1B receptor in mediating the actions of cocaine, we used two model systems: knockout (KO) mice lacking the 5-HT1B receptor and an acute treatment with the 5-HT1B/1D antagonist GR127935. GR127935 attenuated the ability of cocaine to stimulate locomotion and induce c-fos expression in the striatum. However, GR127935 had no apparent effect on the rewarding or sensitizing effects of cocaine. In contrast, as demonstrated previously, the 5-HT1B receptor KO mice showed a heightened locomotor response to cocaine, as well as an increased propensity to self-administer cocaine. Thus, an acute pharmacological blockade of the 5-HT1B receptor decreases some effects of cocaine, while a constitutive genetic KO of the same receptor has opposite effects. These results suggest that compensatory changes have taken place during the development of the 5-HT1B KO mice, which may have rendered these mice more vulnerable to cocaine. The 5-HT1B KO mice should therefore be considered as a genetic model of vulnerability to drug abuse rather than a classic pharmacological tool.

Genetic analysis of the relationships between behavioral and neuroendocrine traits in Roman High and Low Avoidance rat lines.

In order to determine whether the coselection observed between the selection trait (active avoidance behavior) of the Roman High Avoidance (RHA) and Roman Low Avoidance (RLA) rat lines and their neuroendocrine characteristics were genetically determined, we analyzed, in nonsegregating (RHA, RLA, and F1) and segregating (F2 and the two backcrosses) crosses, the inheritance pattern and the phenotypic correlations among behavioral (shuttle-box behavior), physiological (body, adrenal, and thymus weights), and neuroendocrine (corticosterone and prolactin reactivity, catecholamine enzyme activities) variables. Physiological characteristics and enzyme activities have a crucial role in sex dissociation. Avoidance behavior and prolactin reactivity to novel environment remained associated in segregating crosses despite gene rearrangement. They represented the most important variables to differentiate the Roman lines, perhaps sharing a common regulatory mechanism under genetic control.

Psychoneuroendocrine profile associated with hypertension or hyperactivity in spontaneously hypertensive rats.

The behavioral and neuroendocrine reactivity to a novel environment (open field) and the adrenocorticotropic hormone (ACTH)/corticosterone response to a corticotropin-releasing factor (CRF) challenge were measured in 2-mo-old rats from four inbred strains derived from the Wistar-Kyoto rat: spontaneously hypertensive rats (SHRs), hypertensive and behaviorally hyperactive to novelty; WKY, neither hypertensive nor hyperactive; WKHA, hyperactive but normotensive; and WKHT, only hypertensive. The ACTH response to CRF was much lower in SHRs than WKYs, this reduced reactivity being clearly associated with the hyperactivity trait, since it was present in the WKHA and absent in the WKHT strain. On the other hand, the ACTH/corticosterone response to a psychological stimulus (open field) could not clearly discriminate the four strains. The largest difference was found in the prolactin response. Post-open-field levels were much lower in the WKHA (27.11 +/- 4.69 ng/ml) than in the parent WKY strain (83.65 +/- 6.84 ng/ml), the hypertensive strains having intermediate levels (WKHT: 58.05 +/- 7.65 ng/ml; SHR: 64.13 +/- 7.19 ng/ml). Other differences were also found in the levels of aldosterone and renin activity. These results indicate that these strains are an excellent model to study neuroendocrine correlates of hypertension and hyperactivity, which are associated in the SHR strain and may be of interest for the study of the association between neuroendocrine and behavioral characteristics.

5-Carboxamido-tryptamine, CP-122,288 and dihydroergotamine but not sumatriptan, CP-93,129, and serotonin-5-O-carboxymethyl-glycyl -tyrosinamide block dural plasma protein extravasation in knockout mice that lack 5-hydroxytryptamine1B receptors.

We studied the dural plasma protein extravasation response after unilateral electrical stimulation of the trigeminal ganglion in mice lacking serotonin 5-HT1B (5-HT1D beta) receptors by modifying a technique previously described in rats or guinea pigs. We investigated the inhibitory effects of six 5-HT1 receptor agonists in this model: 3-(1,2,5,6-tetrahydropyrid-4-yl)pyrrolo[3,2-b]pyrid-5-one (CP-93,129), sumatriptan, serotonin-5-O-carboxymethyl-glycyl -tyrosinamide (GTI), 5-methylaminosulfonylmethyl-3-(N-methylpyrrolidin-2R -ylmethyl)-1H-indole (CP-122,288), 5-carboxamido-tryptamine (5-CT), and dihydroergotamine. The plasma extravasation response did not differ between wild-type and mutant after vehicle injection. The potency of sumatriptan, CP-122,288, CP-93,129, and 5-CT in wild-type mice was similar to that previously reported for rats. CP-122,288 (1 nmol kg), 5-CT (1 nmol/kg), and dihydroergotamine (72 nmol/kg) inhibited plasma protein extravasation within dura mater after electrical trigeminal ganglion stimulation in both wild-type and knockout mice, which suggests that these agonists act predominantly via receptors other than 5-HT1B. Unlike the wild-type mice, CP-93,129 (1.4 mumol/kg), a specific 5-HT1B receptor agonist, had no effect in knockout mice. The same held true for sumatriptan (0.7 mumol/kg) and GTI (0.6 mumol/kg). These results suggest that CP-93,129, sumatriptan, and GTI exert their effects via 5-HT1B (5-HT1D beta) receptors in mice.