The effect of unilateral neurotoxic lesions to serotonin fibres in the medial forebrain bundle on the metabolism of [H]DOPA in the telencephalon of the living rat

We used quantitative autoradiography to measure the contribution of the 5-hydroxytryptamine (5-HT, serotonin) innervation of rat telencephalon to the synthesis of dopamine (DA) from exogenous l-DOPA. One week after stereotaxic infusions of 5,7-dihydroxytryptamine (5,7-DHT, 1.6 μg) into the right medial forebrain bundle (MFB), rats received [³H]DOPA (200 μCi, i.v.), which circulated for 90 min. The specific bindings in vitro of the 5-HT uptake site ligand [³H]citalopram and the DA uptake site ligand [¹²⁵I]RTI-55 were measured in cryostat sections from the prosencephalon. In most structures ipsilateral to the lesion, [³H]citalopram specific binding was substantially reduced (50–90%). In the lateral habenula specific binding declined by only 30–40%, reflecting the presence of a 5-HT pathway deviating from the MFB at the mesencephalic flexure. [¹²⁵I]RTI-55 binding in the basal ganglia was reduced by 50% on the side of the 5,7-DHT lesion, but was unperturbed in rats pretreated with desmethylimipramine (DMI). 5,7-DHT infusions decreased the synthesis of [³H]DA from [³H]DOPA in vivo in the basal ganglia by (40–90%). Pretreatment with DMI protected [³H]DA synthesis in the basal ganglia, but not in the olfactory tubercle and amygdala ipsilateral to the lesion. Whereas the 5-HT innervation does not contribute greatly to [³H]DA synthesis in the basal ganglia, a substantial proportion of [³H]DA synthesis in olfactory tubercle and amygdala requires an intact 5-HT innervation.     

Ipsilateral alterations in tryptophan hydroxylase activity in rat brain after hypothalamic 5,7-di-hydroxytryptamine lesion

The in vivo relationship between the amounts of tryptophan hydroxylase (TPH) protein and its intrinsic synthetic activity, measured by quantifying the amounts of α-[³H]methyl-5-hydroxytryptamine (α-[³H]M5-HT), is reported in cell body and terminal areas of intact and disturbed serotonergic neurons following a unilateral 5,7-dihydroxytryptamine (5,7-DHT) lesion of the dorsolateral hypothalamus. Five days after the lesion, the relationships between TPH and its synthetic product 5-HT were evaluated on adjacent brain sections in serotonergic cells bodies of the dorsal raphe nucleus (DRN) and nerve fibres of the medial forebrain bundle (MFB). On the side contralateral to the lesion, TPH and α-[³H]M5-HT levels in the intact hemi-DRN exhibited a caudo-rostral distribution and were positively and significantly correlated (P 0.001); the calculated TPH-specific activity was 0.76 nCi of α-[³H]M5-HT formed per U TPH. In the MFB, quantitative measurements of TPH and α-[³H]M5-HT showed no correlation between enzyme and product and no specific activity for TPH could be determined. On the side ipsilateral to the lesion, the density of TPH-immunoreactive fibers was drastically decreased in the dorsolateral hypothalamus where a significant reduction in TPH content (45.5% of control side,P < 0.001) was found. In the overall ipsilateral hemi-DRN, TPH and α-[³H]M5-HT levels, their correlation as well as TPH-specific activity were unaltered by the lesion but a significant increase in α-[³H]M5-HT and TPH contents was observed in the lateral wings of the DRN. The lesion also induced a significant increase in α-[³H]M5-HT and TPH levels (136% and 93.8%,P < 0.001, respectively) in the ipsilateral MFB, which resulted in a positive and significant correlation between these two markers and yielded a TPH-specific activity of 1.0 nCi of α-[³H]M5-HT formed per U TPH. TPH topological area was also significantly increased in the lateral aspect of the ipsilateral MFB 5 days post lesion. These results show that 5-HT synthesis in the intact DRN is proportional to and dependent on TPH activity while in the MFB, 5-HT accumulation appears unrelated to TPH content which is most likely in an inactive enzymatic form. Moreover, the data show that a local disruption of serotonergic terminals in the dorsolateral hypothalamus does not affect 5-HT synthesis in the overall ipsilateral DRN neurons but results in local activation of TPH within the serotonergic projection neurons and the ipsilateral MFB, as evidenced by active de novo synthesis of 5-HT. Altogether the results point to circumscribed activation of compensatory mechanisms in 5-HT synthesis after selective destruction of serotonergic terminals.     

Hippocampal serotonin re-uptake and nocturnal locomotor activity after microinjections of 5,7-DHT in the fornix-fimbria

The role of the hippocampal 5-hydroxytryptamine (5-HT) terminals in the control of locomotor activity was investigated by lesioning 5-HT axons in the fimbria with 5,7-dihydroxytryptamine (5,7-DHT). Rats pretreated with desimipramine (10 mg/kg, i.p.) received microinjections of 5,7-DHT (0, 1, 3, 5 or 10 μg in 0.4 μl ascorbic Ringer's solution) into the fornix-fimbria. On the fourteenth to twenty-first nights after operation, nocturnal locomotor activity was measured in photocell cages. Twenty-eight to thirty days after operation degeneration of 5-HT terminals was assessed by measuring in vitro [³H]5-HT re-uptake in slices of dorsal hippocampus, ventral hippocampus and the septum.Groups injected with 5,7-DHT showed hyperactivity in the night period and increased decrements of activity between tests, both of which were related to the dose of neurotoxin. A reduction of [³H]5-HT re-uptake was found in dorsal hippocampus which was related to the dose of 5,7-DHT, but ventral hippocampal and septal [³H]5-HT re-uptake were not systematically reduced. For each rat, levels of dorsal and ventral hippocampal [³H]5-HT re-uptake were negatively correlated with the mean nocturnal activity from the 7 nights of testing. Levels of dorsal, but not ventral hippocampal [³H]5-HT re-uptake were negatively correlated with the mean nightly decrement of activity. No correlations were found between septal [³H]5-HT and these activity measures. These results, indicate that the increase in nocturnal locomotor activity caused by generalized depletion of 5-HT in the brain may be due to disruption of hippocampal 5-HT terminals supplied by the fornix-fimbria.     

Temporal effects of intrahypothalamic 5,7-dihydroxytryptamine: relationship between serotonin levels and [H]serotonin binding

The relationship between serotonin (5-HT) levels and [³H]5-HT binding in discrete hypothalamic areas was examined in separate groups of animals at various times, following unilateral intrahypothalamic injection of 5,7-dihydroxytryptamine (5,7-DHT). Seven days post-5,7-DHT lesion, 5-HT levels were significantly decreased in both the ipsilateral and contralateral ventromedial and dorsomedial hypothalamic nuclei (VMN, DMN). In the lateral hypothalamic area (LHA), 5-HT levels were significantly decreased only ipsilaterally. Fifty days postlesion, 5-HT levels in the ipsilateral VMN remained significantly below sham, while the DMN and LHA returned to sham values. Seven days after 5,7-DHT there was a significant increase in [³H]5-HT labeling densities in the ipsilateral and contralateral ventromedial hypothalamic area as well as in the ipsilateral LHA. In contrast, in the dorsomedial hypothalamic area there was no increase in [³H]5-HT binding. Fifty days postlesion, no significant differences in [³H]5-HT binding between 5,7-DHT and sham were observed in any areas examined. This data provides further evidence for the regeneration of 5-HT fibers in the hypothalamus and demonstrates that the relationship between [³H]5-HT binding and 5-HT levels varies from one hypothalamic area to another.     

Intrahypothalamic 5,7-dihydroxytryptamine facilitates feminine sexual behavior and decreases [H]imipramine binding and 5-HT uptake

5,7-Dihydroxytryptamine (5,7-DHT) injected into the hypothalamus facilitated feminine sexual behavior in ovariectomized, estrogen-treated female rats beginning 9 days post-lesion. 5,7-DHT treatment was associated with decreased [³H]5-HT but not [³H]NE uptake in the whole hypothalamus and with decreased [³H]-imipramine binding in some hypothalamic nuclei. These data provide the first demonstration using chemical lesions that 5-HT neurons may exert tonic inhibition on hormone-mediated feminine sexual behavior.     

Effects of 5,6-dihydroxytryptamine on nerve terminal serotonin and serotonin uptake in the rat brain

One intraventricular injection of 5,6-dihydroxytryptamine (5,6-DHT) caused the disappearance of fluorescent histochemically detectable 5-hydroxytryptamine (5-HT)-containing terminals and a loss in 5-HT uptake sites. There was an almost complete disappearance of 5-HT-containing nerve terminals in periventricularly located diencephalic areas and in the spinal cord 10–15 days after 75 μg of 5,6-DHT. The noradrenaline and dopamine innervation patterns in the hypothalamus, septum, basal ganglia, and spinal cord appeared normal, except in a narrow zone of the caudate nuclei facing the lateral ventricles, where there was a marked reduction in dopamine fluorescence. These changes were accompanied by 50–87% reductions in the uptake of [³H]5-HT by thin slices of cortex, hypothalamus and spinal cordin vitro. In contrast, the uptake ofL-[³H]noradrenaline was close to normal in hypothalamus and spinal cord slices, and about 35% reduced in the cortex slices. These results are consistent with the idea that intraventricularly administered 5,6-DHT causes extensive axonal degeneration of central serotonin neurones, and that noradrenaline and dopamine neurones are largely unaffected after one injection of 75 μg.     

Sexual behavior in male rats after intracerebral injection of 5,7-dihydroxytryptamine

Adult intact, or castrated testosterone propionate (TP, 150 μg/kg) treated male rats, were tested for masculine sexual behavior after having been injected with 5,7-dihydroxytryptamine (5,7-DHT, 4 μg/4 ml) intracerebrally either alone or in combination with systemic treatment with protriptyline, a noradrenaline (NA) re-uptake blocking agent. No changes were found in the sexual behavior of intact rats although the brain 5-HT levels were reduced to about one-third of their normal value. By contrast, there was a marked increase in the proportion of rats showing ejaculation patterns in the castrate + TP group after 5,7-DHT lesion than in the vehicle-injected group.Compared to the control group, the 5,7-DHT group showed a reduced uptake of [³H]5-HT and [³H]NA in the hypothalamus. Also the uptake of [³H]amines in the cerebral cortex was lowered although the difference did not attain statistical significance. A statistically significant relationship was found between the behavioral changes and the reduction of [³H]5-HT uptake in the hypothalamus while no such relationship was found between the NA uptake and the behavioral changes.Tistochemical analysis of the site of the 5,7-DHT injections showed that the unspecific damage (nerve cell loss, glial cell infiltration) involved a somewhat larger area in the 5,7-DHT group than in the controls. These unspecific lesions were, however, located outside the region of the large medial 5-HT bundle.The results support the hypothesis that 5-HT serves as a transmitter in the neural processes underlying masculine sexual behavior and, further, points to one component of the ascending 5-HT projections which innervates inter alia the hypothalamus as being of particular importance in this context.     

[I]Epibatidine-labelled nicotinic receptors in the extended striatum and cerebral cortex: lack of association with serotonergic afferents

In rat extended striatum, most nicotinic cholinoceptors are likely to be presynaptic. A previous report suggested that DA and 5-HT afferents each account for at least 30% of nicotinic binding sites in the striatum. To explore this question further, rats received unilateral infusions of the neurotoxins 5,7-dihydroxytryptamine, 6-hydroxydopamine or vehicle into the medial forebrain bundle, and were sacrificed 3 weeks later. Denervation was quantified by [¹²⁵I]RTI-55 autoradiography, using separate assay conditions that revealed DA and 5-HT transporters (i.e. DAT and SERT). Nicotinic cholinoceptors were quantified by [¹²⁵I]epibatidine autoradiography. Infusion of 6-hydroxydopamine depleted DAT but not SERT labelling in all striatal areas (i.e. caudate-putamen, nucleus accumbens core and shell, olfactory tubercle). The serotonergic neurotoxin 5,7-dihydroxytryptamine depleted SERT and, to a lesser extent, DAT labelling. Both neurotoxins reduced [¹²⁵I]epibatidine binding in striatal areas. Multiple linear regression analysis showed that these reductions in [¹²⁵I]epibatidine binding were entirely associated with loss of DAT rather than SERT. The DAT-associated proportion of total [¹²⁵I]epibatidine binding was 36±2% (caudate-putamen), 28±3% (accumbens core), 27±4% (accumbens shell) and 44±5% (olfactory tubercle). Cortical [¹²⁵I]epibatidine binding was unaltered by 5,7-dihydroxytryptamine lesions that reduced SERT labelling by 46 to 73%. In all brain areas, even small (3.4 to 8.8%) SERT-associated reductions in [¹²⁵I]epibatidine binding would have been detected as statistically significant. In conclusion, we report the failure to detect nAChRs on 5-HT terminals in extended striatum or cerebral cortex, using a sensitive [¹²⁵I]epibatidine autoradiographic assay.     

Regional central serotonin-2 receptor binding and phosphoinositide turnover in rats with 5,7-dihydroxytryptamine lesions

"Denervation supersensitivity" of serotonin (5-HT) receptors has been proposed to explain the behavioral supersensitivity to 5-hydroxytryptophan (5-HTP) which develops after lesions of indoleamine neurons with 5,7-dihydroxytryptamine (5,7-DHT). To examine the possible role of receptor recognition sites and second messenger activity in supersensitivity, we measured regional 5-HT2 receptor ligand binding and 5-HT-stimulated phosphoinositide turnover in adult rats with 5,7-DHT lesions made by intracisternal injection and their saline-treated controls. In [3H]ketanserin binding studies of fresh brain tissue two weeks after 5,7-DHT injection, there were no significant changes in frontal cortex, brainstem, or spinal cord in Bmax, Kd, or nH of 5-HT2 receptors, 5,7-DHT lesions did not affect basal levels of [3H]inositol phosphate (IP) accumulation but significantly increased 5-HT-stimulated [3H]IP accumulation in the brainstem (+27%) and cortex (+23%). Because brainstem rather than cortex is involved in 5-HTP-evoked myoclonus, increased 5-HT-stimulated phosphoinositide hydrolysis in brainstem following 5,7-DHT lesions in the rat may be relevant to serotonergic behavioral supersensitivity.     

Regeneration of serotonergic fibers in the rat hypothalamus following unilateral 5,7-dihydroxytryptamine injection

The time course of degeneration and regeneration of serotonin (5-HT) fibers in the rat hypothalamus was studied with 5-HT immunocytochemistry and [3H]5-HT uptake following unilateral injections of 5,7-dihydroxytryptamine (5,7-DHT) into the dorsolateral hypothalamus. Within 3 days of the lesion, 5-HT fibers in the ipsilateral hypothalamus were swollen and darkly stained for 5-HT. In the contralateral hypothalamus few swollen fibers were apparent and these were generally restricted to the area adjacent to the fornix. Swollen 5-HT fibers were evident in the ipsilateral hypothalamus 3-19 days post-lesion in the medial forebrain bundle (MFB) during which time there was a gradual decrease in their density. In the medial and periventricular areas of the ipsilateral hypothalamus there were essentially no 5-HT fibers 7-30 days post-lesion. Sprouting 5-HT fibers were observed 12-19 days post-lesion. Thirty days post-lesion the density of 5-HT fibers in the MFB appeared normal; however, medial and periventricular areas remained denervated. Fifty days post-lesion there was an apparent bilateral hyperinnervation in the lateral and dorsomedial hypothalamic areas of 5,7-DHT-injected animals as compared to sham-injected animals. The morphological data were paralleled by changes in [3H]5-HT uptake. Seven days post-lesion specific high affinity uptake was reduced to 27% of sham in the ipsilateral hypothalamus and to 53% of sham in the contralateral hypothalamus. By 50 days post-lesion, specific high affinity uptake of [3H]5-HT was 141% of sham in the ipsilateral hypothalamus and 96% of sham in the contralateral hypothalamus.     

Effects of antipsychotic drugs on dopamine and serotonin contents and metabolites, dopamine and serotonin transporters, and serotonin1A receptors

Summary. The effects of neuroleptics have been attributed to dopamine (DA) receptor blockade; however, other neurotransmitters, in particular serotonin (5-HT), have also been implicated. In this study, we examined the effects of clozapine and haloperidol on the distribution of DA and 5-HT transporters, on endogenous DA, 5-HT and their major metabolites, and on 5-HT_1A receptors. Adult male Sprague-Dawley rats were treated with either haloperidol (1 mg/kg/day, i.p.), clozapine (20 mg/kg/day, i.p.) or saline for 21 days, and following 3 days of withdrawal, the brains were removed. Tissue levels of DA and 5-HT and their metabolites were measured by high-performance liquid chromatography in 16 brain regions, while quantitative autoradiography with [¹²⁵I]RTI-121, [³H]citalopram and [³H]8-OH-DPAT was employed to label DA transporters, 5-HT transporters and 5-HT_1A receptors, respectively. After haloperidol, densities of 5-HT transporters were increased in the dorsal insular cortex and in the ventral half of caudal neostriatum, while 5-HT_1A receptors augmented in cingulate cortex but decreased in the entorhinal area. After clozapine, [³H]citalopram labelling was increased in ventral hippocampus, ventral caudal neostriatum and nucleus raphe dorsalis, but decreased in medio-dorsal and latero-dorsal neostriatum as well as in substantia nigra. Binding of [³H]8-OH-DPAT following clozapine was decreased in frontal, parietal, temporal and entorhinal cortices but increased in the CA3 division of Ammon's horn. The changes in 5-HT transporters in nucleus raphe dorsalis and substantia nigra, as well as the 5-HT_1A receptor down-regulations caused by clozapine but not by haloperidol, may explain effects obtained with clozapine and other atypical neuroleptics. There were no modifications in densities of DA transporters, nor of tissue DA levels, after the chronic neuroleptic treatments. The results are in line with previous suggestions that a certain degree of tolerance to neuroleptics develops, in spite of profound D₁ and D₂ receptor changes that persist during the entire chronic treatment with these psychotropic agents. Received September 2, 1997; accepted July 9, 1998     

Basal and stimulated extracellular serotonin concentration in the brain of rats with altered serotonin uptake

We examined the relationship between the density of serotonergic (5-hydroxytryptamine [5-HT]) uptake sites and extracellular 5-HT concentration in the rat brain using microdialysis with two different models, lesions with 5,7-dihydroxytryptamine (50 μg in the dorsal raphe nucleus (DRN) 15 days before) and sublines of rats genetically selected displaying extreme values of platelet 5-HT uptake. Compared to controls, lesioned rats had a reduced cortical concentration of 5-hydroxyindoles (45%), unchanged basal extracellular 5-HT in the DRN and ventral hippocampus (VHPC), and reduced basal 5-hydroxyindoleacetic acid (5-HIAA) concentrations (46%, DRN; 22%, VHPC). Yet the perfusion of 100 mmol/L KCl or 1 μmol/L citalopram elevated dialysate 5-HT significantly more in the DRN and VHPC of controls. In genetically selected rats, platelet 5-HT content and uptake were highly correlated (r² = 0.9145). Baseline dialysate 5-HT (VHPC) was not different between high and low 5-HT rats and from normal Wistar rats. However, KCl or citalopram perfusion increased dialysate 5-HT significantly more in high 5-HT than in low 5-HT rats, and the former displayed a greater in vivo tissue 5-HT recovery. Significant but small differences in the same direction were noted in [³H]citalopram binding in several brain areas, as measured autoradiographically. Thus, basal extracellular 5-HT (but not 5-HIAA) concentrations are largely independent on the density of serotonergic innervation and associated changes in uptake sites. However, marked differences emerge during axonal depolarization or reuptake blockade. The significance of these findings for the treatment of mood disorders in patients with neurological disorders is discussed. Synapse 28:313–321, 1998. © 1998 Wiley-Liss, Inc.     

Temporal effects of intrahypothalamic 5,7-dihydroxytryptamine: relationship between serotonin levels and [3H]serotonin binding

The relationship between serotonin (5-HT) levels and [3H]5-HT binding in discrete hypothalamic areas was examined in separate groups of animals at various times, following unilateral intrahypothalamic injection of 5,7-dihydroxytryptamine (5,7-DHT). Seven days post-5,7-DHT lesion, 5-HT levels were significantly decreased in both the ipsilateral and contralateral ventromedial and dorsomedial hypothalamic nuclei (VMN, DMN). In the lateral hypothalamic area (LHA), 5-HT levels were significantly decreased only ipsilaterally. Fifty days postlesion, 5-HT levels in the ipsilateral VMN remained significantly below sham, while the DMN and LHA returned to sham values. Seven days after 5,7-DHT there was a significant increase in [3H]5-HT labeling densities in the ipsilateral and contralateral ventromedial hypothalamic area as well as in the ipsilateral LHA. In contrast, in the dorsomedial hypothalamic area there was no increase in [3H]5-HT binding. Fifty days postlesion, no significant differences in [3H]5-HT binding between 5,7-DHT and sham were observed in any areas examined. This data provides further evidence for the regeneration of 5-HT fibers in the hypothalamus and demonstrates that the relationship between [3H]5-HT binding and 5-HT levels varies from one hypothalamic area to another.     

Plastic responses of neonatal 5-hydroxytryptamine1B receptors to 5,7-dihydroxytryptamine lesions mapped by quantitative autoradiography

We previously found different effects on behavior, serotonin (5-HT) concentrations, 5-HT uptake sites, and 5-HT_1A binding sites of neonatal 5,7-dihydroxytryptamine (5,7-DHT) lesions depending on the route of 5,7-DHT injection. To study the impact of early lesions on 5-HT_1B sites as putative 5-HT terminal autoreceptors, we labelled them autoradiographically with [³H]5-HT 4 months after intraperitoneal (i.p.) or intracisternal (i.c.) 5,7-DHT injection during the first postnatal week and quantitated specific binding in 22 brain regions. Changes were confined to the subiculum and substantia nigra, regions with the most 5-HT_1B-specific binding and projection areas of structures with high mRNA expression. Both routes of 5,7-DHT injection were associated with increases in specific binding in subiculum (24% for i.p. and 47% for i.c. route). In contrast, there was a 32% increase in specific binding in the substantia nigra in rats with lesions made i.c. but not i.p. No significant differences were found in nucleus accumbens, caudate-putamen or other brain areas. In saturation homogenate binding studies of 5-HT_1B sites using [¹²⁵I]iodocyanopindolol 1 month after i.p. injections, neonatal 5,7-DHT lesions did not significantly alter B_max or K_d in the neocortex, striatum, diencephalon or brainstem. These data indicate the differential effects of the route of neonatal 5,7-DHT injections on plasticity of 5-HT_1B receptor recognition sites and suggest the presence of a subpopulation of post-synaptically located 5-HT_1B sites which increases in response to denervation. The data also suggest that sprouting of 5-HT neurons after neonatal 5,7-DHT lesions does not involve 5-HT_1B sites.     

Changes in neostriatal DA metabolism after carbachol or atropine microinjections into the substantia nigra

Short infusions (10 min) of carbachol or atropine (5 · 10^?4M) were performed unilaterally in the substantia nigra of the rat. Immediately after, neostriatal dopamine (DA) metabolism was investigated within 15 min.Carbachol infusion produced an increase of the neostriatal DA content. In animals pretreated with Ro 4-4602, a synthesis inhibitor of DA, carbachol application did not change the initial accumulation of newly formed [³H]DOPA from intravenously injected [³H]tyrosine. In such treated animals the normal rate of DA disappearance was reduced.Intranigral atropine infusion produced no change in the neostriatal DA concentration. The initial accumulation of newly formed [³H]DOPA from [³H]tyrosine, in Ro 4-4602 pretreated animals was increased. In such treated animals the normal rate of DA disappearance was activated.In conclusion, intranigral application of carbachol or atropine produced immediate and opposite effects on neostriatal DA metabolism. Carbachol inhibited DA utilization. Atropine activated both DA synthesis and utilization.     

Autoradiographic characterization of [3H]imipramine and [3H]citalopram binding in rat and human brain: species differences and relationships to serotonin innervation patterns.

The neuroanatomical distribution of binding sites for [3H]imipramine and [3H]citalopram was assessed by in vitro autoradiography in select regions of the rat and human forebrain. To determine involvement of serotonin-containing terminals in the binding of [3H]imipramine and [3H]citalopram, binding of these compounds was measured in rats after destroying serotonin-containing neurons with 5,7-dihydroxytryptamine (5,7-DHT). Treatment with this neurotoxin decreased serotonin content by 90% and reduced [3H]citalopram binding to a similar extent. These results demonstrate that [3H]citalopram binding is a reliable marker for serotonin-containing terminals. Binding of [3H]imipramine was reduced by only 15-35% after 5,7-DHT treatment. These latter results suggest that only a small fraction of [3H]imipramine binding to brain sections is associated with serotonergic terminals under standard conditions used in autoradiographic studies with the ligand. Dose-response effects of fluoxetine and desipramine on displacement of [3H]imipramine binding in forebrain regions indicate that the ligand labels predominantly high capacity, low affinity binding sites. To determine the utility of the rat brain as a model for [3H]imipramine and [3H]citalopram binding in the human brain, binding of the ligands was compared in human and rat hypothalamus, amygdala, and hippocampus. The pharmacological characteristics of [3H]imipramine and [3H]citalopram binding were similar in the rat and human brain. However, substantial species differences were observed in topographic patterns of [3H]imipramine binding within the hippocampus and hypothalamus. The distribution of [3H]citalopram binding sites within the amygdala and hypothalamus were also strikingly different in rats compared to humans. This work provides the first demonstration that marked species differences exist in the topography of serotonergic innervation and in the distribution of [3H]imipramine binding sites within the rat and human brain regions examined.     

Compensatory responses in the aging hippocampal serotonergic system following neurodegenerative injury with 5,7-dihydroxytryptamine

This study utilized a multidisciplinary approach to examine injury-induced compensatory responses in the aging hippocampal serotonin transporter (5-HTT), a membrane protein implicated in a variety of neurodegenerative disorders. Age-dependent cellular, anatomical, and physiological changes of the 5-HTT were evaluated in female Fischer 344 rats (2 and 17 months) following denervation of the serotonergic afferents (fimbria-fornix and cingulum bundle) to the dorsal hippocampus using the neurotoxicant 5,7-dihydroxytryptamine (5,7-DHT). Seven days following 5,7-DHT administration, a uniform loss of the hippocampal 5-HTT immunoreactivity was observed in both age groups. However, at 21 days 5-HTT immunoreactivity in young 5,7-DHT-treated animals was similar to control levels, indicative of recovery, while older animals exposed to 5,7-DHT did not show recovery of hippocampal 5-HTT expression. 5-HTT binding site density, as determined by quantitative autoradiography ([3H]citalopram), supported the immunohistochemical results by demonstrating a recovery of 5-HTT binding sites in young, but not old animals, at 21 days following the lesion (P < 0.001). Furthermore, cellular electrophysiological function of hippocampal CA1 pyramidal neurons in 3- and 18-month-old F344 rats at 21 days following 5,7-DHT or vehicle treatment were assessed using in vivo microiontophoretic application of serotonin (5-HT). Independent of changes in sensitivity to the inhibitory effects of 5-HT application, the time to recovery of cell firing following application of 5-HT was significantly increased in the 18-month 5,7-DHT group compared to the 18-month vehicle and 3-month 5,7-DHT groups (60 and 59% increases, respectively; P < 0.05). Overall, these series of studies comprise a model which can be used to identify cellular events underlying both the formation of injury-induced compensatory processes in younger animals and the lack thereof with advancing age.     

Localization of GABAA and GABAB receptor subtypes on serotonergic neurons

The effect of selective destruction of serotonin (5-HT)-containing neurons with 5,7-dihydroxytryptamine (5,7-DHT) on [3H] muscimol and (-)-[3H]baclofen binding was investigated in various rat brain regions. Ten days after intracerebroventricular 5,7-DHT, serotonin levels and [3H]imipramine binding were markedly decreased. 5,7-DHT reduced [3H]muscimol binding only in the mesencephalon, and (-)-[3H]baclofen binding was unmodified in all the areas considered. These results suggest that except in the mesencephalon GABA receptors may not be localized on serotonergic nerve terminals.     

Thioperamide, an H3 Receptor Antagonist Prevents [3H]Glucose Uptake in Brain of Adult Rats Lesioned as Neonates with 5,7-Dihydroxytryptamine

As a first attempt at exploring an association between histaminergic and serotoninergic neuronal phenotypes in glucose regulation, the influence of the histamine H₃ receptor antagonist thioperamide on glucose uptake by brain was determined in rats in which the serotoninergic innervations of brain was largely destroyed perinatally. Male Wistar rats were initially treated on the 3rd day after birth with the serotoninergic neurotoxin 5,7-dihydroxytryptamine (5,7-DHT) (75 μg icv) or saline vehicle (10 μl icv). At 8 weeks lesioned and control rats were terminated in order to validate the effectiveness of 5,7-DHT: reduction in 5-HT and 5-HIAA by 83–91% and 69–83% in striatum, frontal cortex, and hippocampus (HPLC/ED method). Other groups of rats were pretreated with thioperamide (5.0 mg/kg ip) or saline vehicle 60 min prior to 6-[³H]-D-glucose (500 μCi/kg ip). Fifteen-min later rats were decapitated and brains were excised and dissected to remove frontal cortex, striatum, hippocampus, thalamus/hypothalamus, pons, and cerebellum. Liquid scintillation spectroscopy was used to determine that [³H]glucose uptake, which was enhanced in 5,7-DHT lesioned rats in cortex (by 88%), hippocampus, thalamus/hypothalamus, pons and cerebellum (each by 47–56%), and in striatum (by 35%). In contrast, thioperamide prevented the enhancement in [³H]glucose uptake in all brain regions of 5,7-DHT neonatally lesioned rats; and [³H]glucose levels were significantly different in all brain regions (except thalamus/hypothalamus) in thioperamide-versus saline-treated rats. These findings indicate a functional association between histaminergic and serotoninergic systems in brain in relation to glucose regulation.     

Noradrenergic innervation of serotonergic neurons in the dorsal raphe: demonstration by electron microscopic autoradiography

In this study, noradrenergic (NE) terminals in the dorsal raphe were identified by [3H]NE electron microscopic (EM) autoradiography. Lesioning of NE terminals by treatment with the selective catecholamine neurotoxin, 6-hydroxydopamine produced a marked decrease in NE-labelled terminals. [3H]5-HT EM autoradiography of the dorsal raphe produced labelling of cell bodies, dendrites and axons but labelled terminals with synaptic junctions were not observed. Serotonergic (5-HT) neurons were identified at an early stage of degeneration following treatment with the selective 5-HT neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT). When both [3H]NE autoradiography and 5,7-DHT lesioning were combined, a majority of NE-labelled terminals, which formed synaptic specializations, innervated degenerating dendrites. These findings suggest that NE terminals directly innervate 5-HT cells in the dorsal raphe.