M.I.K. Fekete

Noradrenergic innervation of the rat hypothalamus: Experimental biochemical and electron microscopic studies

The concentrations of noradrenaline in individual hypothalamic nuclei and in the median eminence were measured 7-10 days following surgical transections of the lower brain stem or electrolytic lesions of the medullary noradrenaline-containing cell groups. Terminal degeneration in the hypothalamus was studied after the same surgical procedures. Direct, monosynaptic connections between the dorsomedial hypothalamic nucleus and all the noradrenaline-containing cell groups investigated were found. Degenerated synaptic boutons were demonstrated in the median eminence, arcuate, dorsomedial, ventromedial, periventricular and paraventricular nuclei following lesions of the solitary tract and of the lateral reticular nucleus. Biochemical measurements indicate that the pontine-medullary noradrenergic cell groups are the source of hypothalamic norepinephrine. Ascending noradrenergic fibers destined to terminate in the hypothalamus are provided by several cell groups, though the bulk of the NA-fibers seem to originate in the A1-group, in the ventrolateral part of the medullary reticular formation. Most of these fibers join the ventral NA bundle, fewer join the dorsal periventricular tract and several probably also join the dorsal NA bundle. A significant overlap was found in the hypothalamic arborization of the noradrenergic fibers, so that no strict topographical organization seems to be present either in their origin or their termination.

Distribution of norepinephrine and dopamine in cerebral cortical areas of the rat

Concentrations of norepinephrine and dopamine were determined using enzyme isotope assay in 27 microdissected cerebral cortical areas of the rat. A detailed map is presented for microdissection of rat cerebral cortex. Norepinephrine was found in low but still measurable quantities throughout the cortex. Differences between cortical areas are also low. Relatively highest levels were demonstrated in the pyriform, insular and entorhinal cortices. The distribution of dopamine was found to be uneven with a maximal regional difference of 1:24. Concentration of dopamine was in all areas lower than that of norepinephrine. The highest dopamine concentration (2,4 ng/mg protein) was measured in the rostral pyriform cortex but other mesocortical (cingulate, frontal, insular and entorhinal) dopaminergic areas also contained relatively high amounts. Except for the caudal occipital and caudal entorhinal cortices all regions studied contained measurable quantities of dopamine. Its low concentration relative to norepinephrine (below 15%) suggests that in the cortical areas studied dopamine is present as the precursor of norepinephrine.

Salsolinol is a Putative Endogenous Neuro-intermediate Lobe Prolactin-Releasing Factor

The isolation and identification of a prolactin‐releasing factor (PRF) from the neuro‐intermediate lobe of the pituitary gland has been pursued for over a decade. Using high‐pressure liquid chromatography with electrochemical detection (HPLC‐ECD) and gas chromatography/mass spectrometry (GC/MS) (R)‐salsolinol (SAL) (a dopamine‐related stereo‐specific tetrahydroisoquinoline) was found to be present in neuro‐intermediate lobe as well as median eminence extracts of male, intact‐, and ovariectomized female rats. Moreover, analysis of SAL concentrations in neuro‐intermediate lobe revealed parallel increases with plasma prolactin in lactating rats exposed to a brief (10 min) suckling stimulus following 4‐h separation. SAL appears to be a selective and potent stimulator of prolactin secretion in vivo and it was without effect on the secretion of other pituitary hormones. We have also found that SAL can elevate prolactin release, although to a lesser extent, in pituitary cell cultures as well as in hypophysectomized rats bearing anterior lobe transplants under the kidney capsule. Lack of interference of SAL with [3H]‐spiperone binding to AP homogenates indicates that SAL does not act at the dopamine D2 receptor. Moreover, [3H]‐SAL binds specifically to homogenate of AL as well as neuro‐intermediate lobe obtained from lactating rats. Taken together, our data clearly suggest that SAL is synthesized in situ and this compound can play a role in the regulation of pituitary prolactin secretion.

Simultaneous radioenzymatic assay of catecholamines and dihydroxy-phenylacetic acid /DOPAC/; comparison of the effects of drugs on the tuberoinfundibular and striatal dopamine metabolism and on plasma prolactin level

Abstract A simultaneous radioenzymatic assay for catecholamines and DOPAC has been developed. 3 H-Methoxy amines and 3 H-homovanillic acid formed in the presence of 3 H-S-adenosyl-methionine and catechol-O-methyl transferase were separated by organic solvent extraction and chromatography. Chlorpromazine, haloperidol, but not sulpiride increased the DOPAC content in the striatum, without affecting the DOPAC level in the median eminence. γ-Butyrolactone increased the dopamine content also only in the striatum. The monoamine oxidase inhibitor pargyline decreased the DOPAC level in both areas examined. All the substances employed increased plasma prolactin level. The biochemical responses of nigrostriatal and tubero-infundibular dopaminergic neurons may not be analogous.

Binding site of salsolinol: its properties in different regions of the brain and the pituitary gland of the rat

It has been recently shown that salsolinol (SAL) is present in the hypothalamic neuroendocrine dopaminergic (NEDA) system and appears to be a selective and potent stimulator of prolactin (PRL) secretion in the rat. Furthermore, the lack of interference of SAL with 3H-spiperone binding in the striatum and the anterior lobe (AL) of the pituitary gland has been also demonstrated. These data clearly indicate that SAL does not act at the dopamine (DA) D(2) receptors, and suggest that SAL supposedly has a binding site through which the secretion of PRL may be affected. Therefore, binding of 3H-SAL to different regions of the central nervous system (CNS) has been investigated. Specific and saturable binding has been detected in the striatum, cortex, median eminence and in the hypothalamus as well as in the AL and the neuro-intermediate lobe (NIL) of the pituitary gland. K(D) values of the bindings were in the nanomolar range in all tissue tested. 3H-SAL displacing activity of several agonists and antagonists of known DA receptors have also been tested. It has been found that DA and in a lesser extent, apomorphine could displace 3H-SAL, but other DA receptor specific ligands have not been able to affect it. Furthermore, several pharmacologically active compounds, selected on the basis of their influence on DA synthesis, transport mechanisms and signal transduction, have also been tested. Neither mazindol (a selective DA transporter inhibitor) nor clonidine (an alpha(2)-adrenoreceptor agonist) could alter SAL binding. At the same time, L-dopa, carbidopa, benserazide and alpha-methyldopa were able to displace 3H-SAL. The possible changes in SAL binding due to physiological and pharmacological stimuli, like suckling stimulus and reserpine pretreatment (that blocks vesicular monoamine transport in DA terminals), respectively, have also been investigated. In the NIL of the pituitary gland and in the median eminence of the hypothalamus the binding decreased following 10 min of suckling stimulus compared to the binding detected in the same tissues obtained from mothers separated from their pups for 4h and not allowed to be suckled. At the same time, there were no changes in the binding at the AL and striatum. Following reserpine pretreatment that has completely prevented PRL releasing effect of SAL, the binding was significantly augmented. These results support our assumption that SAL should have specific binding sites through which it can affect PRL secretion. Furthermore, it clearly suggests that it may regulate DAergic neurotransmission of NEDA neurons by an altered intracellular or intraterminal synthesis and/or distribution of hypophysiotropic DA.

Adrenergic innervation of the rat hypothalamus

The adrenergic innervation of the hypothalamus was studied by measuring hypothalamic adrenaline levels following surgical transection of the lower brain stem or electrolytic lesion of the medullary adrenaline-containing cell groups. The adrenaline levels in some hypothalamic nuclei and in the median eminence showed a slight decrease after partial transection of the medulla oblongata, whilst there was a pronounced decrease (by 59-78%) 7-10 days following total hemisection or unilateral lesion of the C1-catecholaminergic cell group in the medulla oblongata.

Total and partial hypothalamic deafferentations for topographical identification of catecholaminergic innervations of certain preoptic and hypothalamic nuclei

After total deafferentation of the medial hypothalamus in the rat, noradrenaline concentration in the nucleus arcuatus and the eminentia mediana diminished to one-third. The same decrease of noradrenaline concentration resulted also from partial (anterolateral) deafferentation. Rostral and caudal cuts from the medial hypothalamus did not induce any change in the noradrenaline concentration of the two above-mentioned areas. This indicates that noradrenaline containing axons enter the medial basal hypothalamus laterally from the medial forebrain bundle. Total or partial deafferentation of themedial hypothalamus did not affect noradrenaline or dopamine concentrations in the nucleus preopticus medialis, the nucleus interstitialis striae terminalis, the nucleus hypothalamicus anterior and nucleus supraopticus. The catecholaminergic fibres supplying these regions do not pass the medial hypothalamus, but probably ascend laterally from it, in the medial forebrain bundle. The noradrenaline innervation of the nucleus dorsomedialis takes its origin in the ventral noradrenaline bundle and the fibres from the medial forebrain bundle ascend into the nucleus from the lateral side. After total or lateral deafferentation of the medial hypothalamus, that ransects the fibres running to the nucleus laterally, noradrenaline concentration decreases, apart from the nucleus located within or outside the deafferented island. In this case anterior or posterior deafferentation of the hypothalamus is ineffective. Total deafferentation did not change dopamine concentration in the nuclei of the medial basal hypothalamus, thereby furnishing evidence for its intrahypothalamic origin from the A12 cell group. However, after total deafferentation, some slight decrease of dopamine concentration could be observed in the median eminence. This suggests that the dopamine concentration in the median eminence does not originate exclusively from thenucleus arcuatus but to some extent originates from extrahypothalamic sites. After posterior deafferentation, which destroys the fibres of the incertohypothalamic dopamine system, dopamine concentration in the nucleus dorsomedialis decreases. After total deafferentation of the medial hypothalamus, which isolates the nucleus from the ventral nuclei (and so also from the A12 cell group) of the medial hypothalamus, the dopamine concentration in the nucleus dorsomedialis did not change.

Stress‐ as Well as Suckling‐Induced Prolactin Release is Blocked by a Structural Analogue of the Putative Hypophysiotrophic Prolactin‐Releasing Factor, Salsolinol

Prolactin is secreted from the anterior lobe of the pituitary gland in response both to suckling and to stress. We recently observed that 1‐methyl‐6,7‐dihydroxy‐1,2,3,4‐tetrahydroisoquinoline (salsolinol), produced in the neurointermediate lobe of the pituitary gland, as well as in the medial basal hypothalamus, can selectively release prolactin from the anterior pituitary. Therefore, it has been proposed that salsolinol is a putative endogenous prolactin‐releasing factor (PRF). Here, we report that one structural analogue of salsolinol, 1‐methyl‐3,4‐dihydroisoquinoline (1MeDIQ), can block salsolinol‐induced release of prolactin, but does not affect prolactin release in response to thyrotropin releasing hormone (TRH), α‐methyl‐p‐tyrosine (αMpT) (an inhibitor of tyrosine hydroxylase), domperidone (a D2 dopamine receptor antagonist), or 5‐hydroxytryptophan (5‐HTP), a precursor of serotonin). 1MeDIQ profoundly inhibited suckling‐, immobilization‐, as well as formalin‐stress induced prolactin release without any influence on corticosterone secretion. The 1MeDIQ‐induced reduction in prolactin response to immobilization stress was dose‐dependent. These results suggest that salsolinol can play a pivotal role in the regulation of prolactin release induced by either physiological (suckling) or environmental (stress) stimuli.

Anxiolytic 2,3-benzodiazepines, their specific binding to the basal ganglia.

Over the past 20 years, several members of the 2,3-benzodiazepine family have been synthesized. Some of these compounds--tofisopam (Grandaxin), girisopam, nerisopam--exert significant anxiolytic and antipsychotic activities. Sites where actions of 2,3-benzodiazepines are mediated differ from those of 1,4-benzodiazepines. Binding of 2,3-benzodiazepines to neuronal cells in the central nervous system shows a unique and specific distribution pattern: their binding sites are located exclusively to the basal ganglia. Chemical lesioning of the striato-pallido-nigral system, surgical transections of the striato nigral pathway and the activation of c-fos expression in the basal ganglia after application of 2,3-benzodiazepines suggest that these compounds mainly bind to projecting neurons of the striatum. The binding sites are transported from the striatum to the substantia nigra and the entopeduncular nucleus. Recent studies on mechanism of action of 2,3-benzodiazepines indicate their possible role in opioid signal transduction since 2,3-benzodiazepines augment the agonist potency of morphine to induce catalepsy and analgesia, and their action is diminished in morphine tolerant animals. The possible biochemical target of 2,3-benzodiazepines is an alteration in the phosphorylation of protein(s) important in the signal transduction process. Agents affecting emotional responses evoked by endogenous opioids without danger of tolerance and dependence may represent a new therapeutic tool in the treatment of addiction and affective disorders.

Dopamine, noradrenaline and 3,4-dihydroxyphenylacetic acid (DOPAC) levels of individual brain nuclei, effects of haloperidol and pargyline

Noradrenaline (NA), dopamine (DA) and DOPAC were determined with a newly developed radioenzymatic method simultaneously in the striatum, limbic system, hypothalamus and in catecholamine-containing cell groups of the rat brain. Only a loose relationship could be established between DOPAC and DA contents in the various brain areas. The lowest relative DOPAC level (DOPAC/DA ratio) was found in the median eminence, while it was the highest in the periventricular nucleus of the hypothalamus. Haloperidol increased the DOPAC level in only part of the nuclei examined (striatum, olfactory tubercle, central amygdaloid nucleus), while in other limbic regions as well as in the hypothalamic dorsomedial, arcuate and paraventricular nuclei it proved to be ineffective. The DOPAC level in the locus coeruleus was decreased by haloperidol. Pargyline caused an appr. 50% decrease of DOPAC content of most of the nuclei in 10 min; the effectivity of the drug did not show parallelism with that of haloperidol. The monoamine oxidase inhibition caused no change in the DOPAC level in the hypothalamic periventricular and paraventricular nuclei. Results are discussed as a consequence of different reactivity of various DA-ergic terminals and catecholamine cell bodies to haloperidol and pargyline.