Evidence for the existence of serotonin-, dopamine-, and noradrenaline-containing neurons in the gut of Lampetra fluviatilis

Summary Monoamine-containing neurons in the gut of Lampetra fluviatilis are characterized by histochemical, electron microscopical and biochemical methods. Strongly yellow fluorescent, probably serotonin-containing intrinsic neurons are found along the entire length of the intestine. Their processes aggregate to form large bundles of mainly non-terminal axons, constituting a subepithelial fibre plexus. This subepithelial, ganglion cell comprising plexus is connected to a wide-meshed subserosal plexus which has ganglion cells of different size and few varicose, single axons. Intermingled with both plexus there occur — in the anterior and middle but not in the preanal portion of the lamprey intestine — scattered green fluorescent intrinsic perikarya, emanating faintly green fluorescent, poorly varicosed axons.The formaldehyde-induced neuronal fluorophores conform to serotonin (yellow fluorescent compound), noradrenaline, and dopamine (green fluorescent substance), as revealed in microspectrofluorimetric recordings. The electron microscopical analysis of the yellow fluorescent intrinsic neurons in the terminal hindgut shows nerve cell pericarya and axons equipped with a typical population of occasional small granular and many large granular vesicles (750–1600 Å). The number and opacity of cores of the small and the osmiophilia of the cores of the large granular vesicles are significantly increased following short-term treatment with 5,6-dihydroxytryptamine. Long-term treatment with 5,6- or 5,7-dihydroxytryptamine provokes severe signs of ultrastructure impairment and eventual degeneration in the supposed serotonin-containing axons, besides indications of piling-up of organelles in the non-terminal axons due to arrest of axonal transport.Chromatography of acid extracts from the lamprey intestine, gills and kidney reveals the presence of serotonin (besides another unidentified indoleamine) and dopamine and noradrenaline in the gut, but only dopamine in the brain. The detection of serotonin, noradrenaline and dopamine in the lamprey gut is confirmed by chemical determinations.The occurrence of intrinsic serotonin-, noradrenaline- and dopamine-containing neurons in the gut of Lampetra fluviatilis deviates from the established pattern of innervation of the vertebrate intestine and is considered to be a remnant of an autonomic innervation principle common in invertebrates.Supported by grants from the Deutsche Forschungsgemeinschaft.Supported by grants from the Swedish Medical Research Council (No. 14X-712 and 14X-56.The authors are indebted to Lilan Bengtsson, Gertrude Stridsberg, Eva Svensson and Rolf Frank for skilful technical assistance.     

Relation between brain 5-HIAA levels and the release of serotonin into brain synapses

Our findings in experiments using reserpine, an amine releaser, and fluoxetine, a serotonin uptake blocker, indicate that the reuptake of serotonin from brain synapses precedes its transformation to 5-hydroxyindoleacetic acid (5-HIAA). Male rats were injected with reserpine or fluoxetine alone, or with fluoxetine one hour before reserpine; control animals received diluents. Reserpine lowered brain serotonin and raised brain 5-HIAA levels. Fluoxetine alone did not change serotonin levels but lowered 5-HIAA. Fluoxetine completely antagonized the reserpine-induced increase in 5-HIAA, and significantly enhanced its depletion of serotonin. In order to determine whether the ability of fluoxetine to block the rise in 5-HIAA after reserpine resulted from its effect on serotonin reuptake or from suppression of impulse flow along serotoninergic neurons, we also examined the effects of the drugs on serotonin metabolism in distal portions of acutely transected neurons (which, presumably, were no longer able to conduct impulses). No differences were noted between the responses of intact and lesioned serotoninergic neurons, indicating that fluoxetine's blockade of the rise in brain 5-HIAA results from its effect on serotonin reuptake.     

Amnesia,memory and brain systems.

Bilateral damage to either the medial temporal lobe or the diencephalic midline causes an amnesic syndrome, i.e. a global impairment in the ability to acquire new memories regardless of sensory modality, and a loss of some memories, especially recent ones, from the period before amnesia began. The memory deficit can occur against a background of intact intellectual and perceptual functions. Two themes have been prominent in recent work. First, the amnesic syndrome is narrower than once believed in the sense that a number of learning and memory abilities are preserved (e.g. skill and habit learning, simple forms of conditioning and the phenomenon of priming). Second, the brain system damaged in amnesia has only a temporary role in memory. As time passes after learning, memory is reorganized and consolidated within neocortex, such that eventually medial temporal lobe and diencephalic structures are not needed for storage or retrieval.     

Interaction between noradrenergic and glucocorticoid brain systems: Probable involvement in the development of depression

Electrophysiological and biochemical experiments on slices of the rat dorsal hippocampus demonstrated that dexamethasone (100 nM) augmented and prolonged the depressive effect of noradrenaline on synaptic transmission in the CA1 zone; this effect is related to weakening of the uptake of noradrenaline by neurons. The effect of dexamethasone is mediated by glucocorticoid receptors. Inhibitors of presynaptic translocase of noradrenaline, cocaine and imipramine, increased, similarly to dexamethasone, the effects of noradrenaline; an additive synergism was observed upon combined applications of dexamethasone and cocaine. The effect of dexamethasone decreased with an increase in the extracellular concentration of glucose, but increased upon application of the Na/K-ATPase inhibitor strophantin. The potentiating influence of dexamethasone on the effects of noradrenaline was weaker in slices obtained from rats with behavioral depression induced by social isolation or chronic introduction of dexamethasone. We hypothesize that glucocorticoids stabilize noradrenergic neurotransmission in the brain under the action of stressogenic influences. The role of glucocorticoid mechanisms in the development of depression is discussed.Neirofiziologiya/Neurophysiology, Vol. 36, Nos. 5/6, pp. 377–385, September–December, 2004.This revised version was published online in April 2005 with a corrected cover date and copyright year.     

Relationship between three intensity levels of Toxocara canis larvae in the brain and effects on exploration, anxiety, learning and memory in the murine host

Outbred LACA mice were administered low (100 ova), medium (1000 ova), high (3000 ova) and trickle (4x250 ova) doses of Toxocara canis ova and the effect of infection was examined with respect to the number of larvae recovered from the brain and their behaviour. Recovery of larvae from the brain was generally low with the % recovery expressed in terms of the total dose administered being highest for the 3000 dose (6.1%) and 1000 dose (6%), followed by the 100 (4.4%) and trickle (3.5%) doses. The variation in larval recoveries was large between individual mice receiving similar doses. The level of infection in the brain was lower in mice receiving a multiple as opposed to an equivalent single dose of ova. Mice were then divided into three larval intensity groupings based upon the number of larvae recovered from their brain. The ranges for the groups were as follows: low intensity group, 0-15 larvae; moderate intensity group, 27-55 larvae; high intensity group, 66-557 larvae. Three behavioural tests were carried out on control and infected mice. Exploration and response to novelty was examined using a 'T' maze and learning was investigated by means of a water-finding task. Anxiety was measured using an elevated plus maze apparatus. Infected mice were less explorative and less responsive to novelty in the 'T' maze and this was particularly pronounced for the heavily infected mice. In the elevated plus maze, infected mice displayed reduced levels of anxiety to aversive and exposed areas of the maze, particularly in the case of the moderate and high intensity mice. There was evidence for impaired learning ability in the water task apparatus for moderate and high intensity mice. In general, the effects of infection on behaviour were more pronounced in the moderate and high intensity groups compared to the low intensity group.     

Difference in susceptibility of arginine-vasopressin and oxytocin to aminopeptidase activity in brain synaptic membranes

Arginine-vasopressin and oxytocin, peptides which serve as putative precursors for neurotrophic fragments, were digested in the presence of the respective ¹⁴C-Tyr²- and ${}^{14}\text{C-GlyNH}{}_2{}^9\text{-labeled}$ nonapeptides with a purified synaptic membrane preparation of rat brain. In this preparation aminopeptidase activity predominates in the conversion of these peptides. The disappearance of intact peptide and the release of free ¹⁴C-Tyr and ¹⁴C-GlyNH₂ was followed simultaneously with time by HPLC. Oxytocin was about four times more resistant to proteolysis than arginine-vasopressin as measured by slower disappearance of intact oxytocin, and reflected by the slower release of ¹⁴C-Tyr, but not of ¹⁴C-GlyNH₂ from oxytocin. Comparison of degradation rates of structure analogues showed that peptides having Ile in position 3, as oxytocin, were more resistant than analogues having Phe in position 3, as arginine-vasopressin. The data demonstrate that arginine-vasopressin and oxytocin differ markedly in susceptibility to the aminopeptidase activity in brain synaptic membranes, and indicate that this difference resides primarily in the amino acid residue in position 3. It is suggested that the difference in susceptibility may affect the pattern of neurotrophic metabolites in brain.     

Central noradrenergic activity and the formation of glycol sulfate metabolites of brain norepinephrine

The present study utilized intraventricular injection of Na₂³⁵SO₄ to detect drug induced changes in the $\text{in vivo}$ formation of the two major metabolites of rat brain norepinephrine (NE) - the sulfate conjugates of 3-methoxy-4-hydroxyphenylglycol (MOPEG-SO₄) and 3,4-dihyd (DOPEG-SO₄). Assays involved the hypothalamus only. Rats pretreated with clonidine showed a reduced formation of both MOPEG-³⁵SO₄ and DOPEG-³⁵SO₄ after intraventricular Na₂³⁵SO₄ as well as reduced synthesis of ³H-NE from intraventricular ³H-tyrosine. Phenoxybenzamine (POB) produced increases in the synthesis of both ³⁵S-labeled conjugates and ³H-NE. Neither drug altered the loss of exogenous ³H-MOPEG-SO₄ but clonidine increased both the accumulation of labeled sulfate and the sulfation of exogenous MOPEG in pheniprazine treated rats. These results show that the rates of formation of the labeled glycol sulfates are sensitive indicators of changes in brain NE turnover but can also be influenced by factors involved in sulfation that are unrelated to NE turnover. Blockade of NE synthesis with alpha methyltyrosine did not affect resting or POB-elevated levels of the labeled conjugates until stores of NE were reduced by 40%. The latter findings suggest that central noradrenergic neurons can release and metabolize NE at a normal rate despite synthesis blockade so long as adequate stores of NE are available.     

Basal, dopamine- and octopamine-stimulated adenylate cyclase activity in the brain of the moth, M amestra configurat A, during its metamorphosis.

—The specific activities of basal, dopamine- and octopamine-stimulated adenylate cyclase in the brain of the moth, Mamestra configurata, increased by an order of magnitude or more during the transition from pupal to adult brain. Increases in adenylate cyclase activity outpaced increases in brain weight and protein content by a wide margin, especially during the last 10 days of metamorphosis when most of the change in enzyme activity occurred. The greatly enhanced adenylate cyclase activity appears to reflect increased demands placed on the adult brain as an integrative centre, especially in the integration of sensory input from the eyes and antennae. Growth of the optic lobes during brain development contributed a substantial proportion of basal (33%), dopamine-sensitive (21%) and octopamine-sensitive (37%) adenylate cyclase activity to the adult brain. High levels of octopamine and the presence of active octopamine-sensitive adenylate cyclase suggest a cyclic AMP-mediated physiological function for octopamine in the optic lobes of the insect brain, probably as a neurotransmitter or neuromodulator.     

Relation of brain regional physostigmine concentration to cholinesterase activity and acetylcholine and choline levels in rat

The relationship between physostigmine (Phy) concentration, acetylcholine (ACh), choline (Ch) and cholinesterase (ChE) activity was examined in whole rat brain after the administration of [³H]Phy (650 g/kg i.m.). Cholinesterase inhibition was found to be inversely related to Phy levels. Maximal inhibition (80%) was seen at 5 min and by 2 hrs ChE activity had returned to control levels. Acetylcholine levels in whole brain peaked at 30 min at a concentration (80 nmol/g) 2.3 times higher than controls (33 nmol/g). Choline levels were not significantly altered. The regional distribution of Phy concentration and ChE activity was studied in six areas of the brain following i.m. administration of three different dosages of [³H]Phy. Physostigmine concentration and ChE activity showed a dose dependency in each area examined except in SP (medial septum). Striatum (ST) showed the greatest relative increase of ACh up to 30 min, when compared to other areas. Choline levels were not changed in any area with the exception of ST at 5 min where a decrease was seen. There was a relationship between ChE activity, Phy concentration and ACh levels in all areas examined with exception of the medulla oblongata (MO). Our results indicate that even though ChE was inhibited practically uniformly in all brain areas, the percent increase with respect to control animals and the relative increase of ACh varied widely from area to area. This finding has clinical implications in cases in which cholinomimetic therapy is used to elevate ACh levels in specific brain areas which show a cholinergic deficit.Special issue dedicated to Prof. Eduardo De Robertis.     

Nature of stress: differential effects on brain acetylcholinesterase activity and memory in rats

Effect of acute, chronic-predictable and chronic-unpredictable stress on memory and acetylcholinesterase (AChE) was investigated in rats. The animals were subjected to 3 type of stressors--(1) acute immobilization stress, (2) chronic-predictable stress i.e., immobilization daily for 5 consecutive days and (3) chronic-unpredictable stress that included reversal of light/dark cycle, over-night fasting, forced-swimming, immobilization and forced exercise in random unpredictable manner daily for 5 consecutive days. Learning and memory function was studied by single trial Passive avoidance test. AChE activity was assayed spectrophotometrically in the detergent (DS) and salt (SS) soluble fractions in different brain regions. Learning was obtained in acute and chronic-predictable stress groups but not in chronic-unpredictable group. Acute, chronic-predictable and chronic-unpredictable stress caused significant decrease in AChE activity in the DS fraction of cortex, hippocampus and hypothalamus as compared to control. Results indicate that AChE in DS fraction is predominantly affected in stressed and stressed-trained group but cognition is affected only by chronic-unpredictable stress. In acute and chronic-predictable groups the decreased AChE activity in the hippocampal DS fraction during learning may be responsible to maintain cognitive function by enhancing the cholinergic activity.     

The organization of serotonin-, dopamine-, and FMRFamide- containing neuronal elements and their possible role in the regulation of spontaneous contraction of the gastrointestinal tract in the snail Helix pomatia

Summary The distribution of serotonin-, tyrosine hydroxylase-, and FMRFamide-immunoreactive neuronal elements, as well as the concentrations of serotonin and dopamine in the different parts of the gastrointestinal tract, were studied in the snail Helix pomatia. The sensitivity of the spontaneous contractions of the alimentary tract to serotonin, dopamine, and FMRFamide was also tested. Serotonin-, tyrosine hydroxylase-, and FMRFamide-immunoreactive elements could be demonstrated in each part of the gastrointestinal tract, but they showed different innervation patterns. Serotonin- and tyrosine hydroxylase-immunoreactive elements were dominant in the submucosal layer, whereas FMRFamide-immunoreactive elements were dominant in both the mucosal and submucosal layers. Tyrosine hydroxylase-immunoreactive elements were confined to the longitudinal muscle trabeculae of submucosa, whereas serotonin-immunoreactive elements were distributed throughout the submucosal layer. No serotonin-immunoreactive cell bodies, but only fibers, could be detected in the gastrointestinal tract, and therefore they represent extrinsic elements. Tyrosine hydroxylase- and FMRFamide-immunoreactive cell bodies represent intrinsic elements of the tract. The occurrence and density of the serotonin- and tyrosine hydroxylase-immunoreactive elements showed significant differences in the different parts of the alimentary tract, in accordance with HPLC assays, which revealed a significant frontocaudal decrease in both the serotonin (from 2.11 to 1.21 pM/mg) and dopamine (from 3.28 to 0.52 pM/mg) contents of the different parts of the alimentary tract. Dopamine at 10-5 M concentration proved to be effective only on the longitudinal muscles by increasing the tone and frequency of contractions, but was ineffective on the circular muscles. Serotonin affected both the longitudinal and circular muscles. Serotonin at 10-5 M concentration decreased the tone and increased the frequency of low-amplitude contractions of the longitudinal muscles of the esophagus and the gizzard but increased both the tone and frequency of the crop. Serotonin at 10-9 M concentration slightly decreased the tone and blocked the contractions of the circular muscles in the crop but at 10-5 M concentration induced contractions of the circular muscles in the gizzard. FMRFamide at 10-6 M concentration decreased the tone and was shown to block the contractions of both the longitudinal and circular muscles.     

Relation between the structure of the skate brain and its ecology

The original and the literature data on the brain structure in 20 species of skates have been examined in connection with their systemic position and ecology. Certain morpho-ecological brain peculiarities have been defined which are also specific for other groups of Chondrostei, such as: sharks and Holocephala. Allunion Institute of Marine Fishery and Oceanography, Moscow.     

Involvement of noradrenergic transmission in the PVN on CREB activation, TORC1 levels, and pituitary-adrenal axis activity during morphine withdrawal

Experimental and clinical findings have shown that administration of adrenoceptor antagonists alleviated different aspects of drug withdrawal and dependence. The present study tested the hypothesis that changes in CREB activation and phosphorylated TORC1 levels in the hypothalamic paraventricular nucleus (PVN) after naloxone-precipitated morphine withdrawal as well as the HPA axis activity arises from α(1)- and/or β-adrenoceptor activation. The effects of morphine dependence and withdrawal on CREB phosphorylation (pCREB), phosphorylated TORC1 (pTORC1), and HPA axis response were measured by Western-blot, immunohistochemistry and radioimmunoassay in rats pretreated with prazosin (α(1)-adrenoceptor antagonist) or propranolol (β-adrenoceptor antagonist). In addition, the effects of morphine withdrawal on MHPG (the main NA metabolite at the central nervous system) and NA content and turnover were evaluated by HPLC. We found an increase in MHPG and NA turnover in morphine-withdrawn rats, which were accompanied by increased pCREB immunoreactivity and plasma corticosterone concentrations. Levels of the inactive form of TORC1 (pTORC1) were decreased during withdrawal. Prazosin but not propranolol blocked the rise in pCREB level and the decrease in pTORC1 immunoreactivity. In addition, the HPA axis response to morphine withdrawal was attenuated in prazosin-pretreated rats. Present results suggest that, during acute morphine withdrawal, NA may control the HPA axis activity through CREB activation at the PVN level. We concluded that the combined increase in CREB phosphorylation and decrease in pTORC1 levels might represent, in part, two of the mechanisms of CREB activation at the PVN during morphine withdrawal.     

Correlation between the levels of calcium in the blood and catecholamines in the brain during memory formation and fixation in hypoparathyreosis

Relationship between the blood level of calcium and the level of catecholamines in the brain limbic structures was studied in passive avoidance conditioning and extinction in rats with hypoparathyreosis. After parathyroidectomy, conditioning processes were shown to be impaired as a result of a disorder of calcium supply. In hypoparathyreosis, not only the basic dopamine and noradrenalin levels change, but catecholamine dynamics in learning and forced extinction of a passive avoidance reaction shifts. The results point to the deranged functioning of dopamine and noradrenaline brain systems as a result of disorders in calcium homeostasis. These shifts result in disorders of conditioning and development of an adaptive behavioral strategy.