GABAA and GABAB receptor-mediated inhibition of release of 5-hydroxytryptamine in the intermediate lobe of the rat pituitary gland.

Isolated neurointermediate lobes or neural lobes of the rat pituitary gland attached to the pituitary stalk were incubated in vitro and the spontaneous or electrically (pituitary stalk stimulation, 5 Hz, 1,500 pulses) evoked release of 5-hydroxytryptamine was determined. The evoked release of 5-hydroxytryptamine from the neurointermediate lobe was increased fivefold in the presence of the dopamine receptor antagonist (-)-sulpiride (1 microM). The evoked release of 5-hydroxytryptamine from the isolated neural lobe was not altered by (-)-sulpiride. The evoked release of 5-hydroxytryptamine from the isolated neural lobe in the presence of (-)-sulpiride was less than 5% of that from the combined neurointermediate lobe showing that most of the 5-hydroxytryptamine released from the combined neurointermediate lobe originated in the intermediate lobe. The evoked release of 5-hydroxytryptamine from the neurointermediate lobe in the presence of (-)-sulpiride showed a diurnal variation. It was three to five times higher between 9.30 and 14.00 h than between 8.30 and 9.30 h or between 14.00 and 16.00 h. The 5-hydroxytryptamine tissue content at the end of the incubation experiments also showed similar variations which were, however, less pronounced. The evoked release of 5-hydroxytryptamine from the neurointermediate lobe, in the presence of (-)-sulpiride, was reduced by the preferential GABAA receptor agonist muscimol or the selective GABAB receptor agonist (-)-baclofen in a concentration-dependent manner. Bicuculline, a selective GABAA receptor antagonist inhibited the effect of muscimol, but not that of (-)-baclofen. Bicuculline alone did not affect the release of 5-hydroxytryptamine from the gland. It is concluded that the release of 5-hydroxytryptamine in the intermediate lobe is influenced by both dopaminergic and GABAergic mechanisms.

Acetylcholinesterase in the rat neurohypophysis is decreased after dehydration and released by stimulation of the pituitary stalk.

Specific acetylcholinesterase and non-specific cholinesterases are present in all three lobes of the rat pituitary gland. This paper describes two new observations on hypophyseal acetylcholinesterase. Firstly, a prolonged increase of neurohormone secretion evoked by dehydration and sodium loading was accompanied by a decrease in the acetylcholinesterase activity localized to the neural lobe, where acetylcholinesterase has previously been demonstrated in fine nerve fibres. Secondly, electrical stimulation of the pituitary stalk in vitro elicited the release of acetylcholinesterase and non-specific cholinesterases from the combined neural and intermediate lobe indicating that the enzyme can be released from nerve endings in the hypophysis by action potentials. The observed loss of enzyme activity during dehydration may be the consequence of a prolonged activation of cholinergic nerves in the gland, leading to an increased release of acetylcholinesterase, which is not immediately replaced by fresh enzyme. The decrease in acetylcholinesterase in the neural lobe during dehydration may also be connected with its peptidase function and thus with the previously observed loss of substance P from the neural lobe during dehydration [Holzbauer et al. (1984) Neurosci. Lett. 47, 23-28].

Variations in the monoamine oxidase-inhibitory activity ("tribulin?") in pig's urine.

Sandler and his colleagues (see Sandler, 1982) have demonstrated the presence of an endogenous inhibitor of the enzyme monoamine oxidase (MAO) and of benzodiazepine receptor binding in the urine and blood plasma of man and rat. The concentrations of this material increased under stress conditions and it has been named "tribulin". In the present experiments MAO-inhibitory activity was found in extracts of urine and plasma samples of domestic pigs. Evidence was obtained that the inhibitory activity was higher when pigs experienced slight discomfort. Thus it appears that pigs produce a substance similar to tribulin. It may become possible to use such MAO-inhibitory activity as an indicator in the assessment of interaction with the environment in pig husbandry.

Release of endogenous 5-hydroxytryptamine from the intermediate lobe of the rat pituitary gland is inhibited by endogenous dopamine.

The release of endogenous 5-hydroxytryptamine (5-HT) from the in vitro incubated neurointermediate lobe (NIL) of the rat hypophysis was measured by HPLC with electrochemical detection. 5-HT release evoked by electrical stimulation of the pituitary stalk from the NIL, but not from the isolated neural lobe (NL) was enhanced in the presence of the dopamine receptor antagonist sulpiride (1 microM). The opiate receptor antagonist naloxone (1 microM) had no effect on the evoked release of 5-HT from the NIL or NL. In conclusion, the release of endogenous 5-HT from the intermediate lobe is under inhibitory control of endogenous dopamine.

Dehydration increases the electrically evoked dopamine release from the neural and intermediate lobes of the rat hypophysis.

The effect of dehydration on dopamine (DA) release from the neural and intermediate lobes of the rat pituitary gland after electrical stimulation of the pituitary stalk was studied in vitro. The amount of DA released from pituitary lobes of dehydrated rats (no water for 72 h), in which the hormone secretion from the hypophysis, especially the neural lobe, is known to be greatly enhanced, was significantly higher when compared with that from control tissues. In control and dehydrated rats the evoked DA release from the intermediate lobe was higher than that from the neural lobe. After dehydration, the stimulus-evoked DA release from the neural lobe increased by more than 200%, whereas that from the intermediate lobe increased by only 60%, indicating a preferential activation of the DA system of the neural lobe. There was a decrease in DA released per pulse from the combined neuro-intermediate lobe of control rats with increasing number of stimulation pulses. In contrast, DA release per pulse from the neuro-intermediate lobe of dehydrated rats showed a tendency to increase with increasing number of pulses. As the availability of newly synthesized DA is believed to be the limiting factor for transmitter release, the present observations suggested a higher in vitro DA synthesis rate in the tissues from dehydrated rats. This agrees with in vivo results of Alper and his colleagues who found an increased accumulation of L-3,4-dihydroxyphenylalanine (DOPA) in neuro-intermediate lobes of dehydrated rats after inhibition of DOPA-decarboxylase.

Monoamines in tubero-hypophyseal structures of farm animals.

Dopamine (DA) and 5-hydroxytryptamine (5-HT) were measured in several regions of the pituitary gland of Jersey steers, Soay ewes and two breeds of pigs. The glands were collected from animals which had not been stressed and the amine concentrations were comparable to those found previously in the pituitary glands of laboratory animals. Metabolites of both amines were present in the infundibulum, the pituitary stalk, the combined neural and intermediate lobe and the anterior lobe of all glands analysed indicating that the amines are being utilized in these structures under "near physiological" conditions.

Release of endogenous 5-hydroxytryptamine from the neural and the intermediate lobe of the rat pituitary gland evoked by electrical stimulation of the pituitary stalk.

A calcium-dependent release of 5-hydroxytryptamine from the neural and intermediate lobe of the rat pituitary gland has been demonstrated following electrical stimulation of the pituitary stalk with stimulation parameters thought to evoke propagated action potentials. The 5-hydroxytryptamine release from the intermediate lobe was double that from the neural lobe. The mass of the intermediate lobe of the rat is about 80% of that of the neural lobe [Holzbauer, Racké, Mann, Cooper, Cohen, Krause and Sharman (1984) J. Neural Transm. 59, 91-104]. The relatively high overflow of 5-hydroxytryptamine from the intermediate lobe agrees with immunohistochemical studies in which a larger number of 5-hydroxytryptamine fibres were seen in the intermediate lobe than in the neural lobe. The present results have demonstrated that the rat hypophysis contains neuronal 5-hydroxytryptamine. They also suggest that this amine may act as a neurotransmitter substance in the neural and intermediate lobe.

The dopaminergic innervation of the intermediate lobe and of the neural lobe of the pituitary gland.

The intermediate and the neural lobe of the pituitary gland are innervated by two, virtually independent, groups of dopaminergic neurons which, until recently, were considered as a uniform system and referred to as the "tubero-hypophyseal dopamine system". Some aspects of the separate physiological functions of these neurons in the intermediate and in the neural lobe, of their microanatomy and biochemistry as well as of dopamine release from their terminals are discussed in this review.

In vivo secretion of 3 alpha-hydroxy-5 alpha-pregnan-20-one, a potent anaesthetic steroid, by the adrenal gland of the rat.

3 alpha OH-5 alpha-Pregnan-20-one (allo-THP), a steroid with strong anaesthetic properties, was found to be secreted by the adrenal gland of the rat in quantities similar to those secreted by the rat ovary. From the hypnotic potencies established for this and other endogenous steroids there can be little doubt that the total amount of steroids with anaesthetic properties produced in a female rat are sufficient to exert a depressant action on certain cells of the brain. In rats with intact adrenal glands a positive correlation existed between the adrenal secretion of allo-THP and pregnenolone or progesterone, whereas that between allo-THP and DOC was negative. This could be the result of a competition between the enzymes responsible for the oxidation and reduction of progesterone, the common precursor of allo-THP and DOC. The possibility that allo-THP could have hypotensive actions was suggested.

Pituitary 5-hydroxytryptamine nerves--a possible link with pituitary hormone secretion.

A short review of the present knowledge on the innervation of the pituitary gland by 5-hydroxytryptamine (5-HT) containing nerves is given. This is followed by results of recent experiments in which the concentrations of 5-HT in different lobes of the rat hypophysis were measured under conditions in which pituitary hormone secretion was increased. A prolonged increase in hormone secretion induced by dehydration and increased sodium chloride intake resulted in a decrease in the 5-HT concentrations in the anterior lobe (AL). The 5-HT content of the neural (NL) and the intermediate lobe (IL) of these rats was hardly changed whereas that of DA was about doubled. Acute stimulation of pituitary hormone secretion by exsanguination under ether anaesthesia also caused a fall in the 5-HT content of the AL. In contrast, in the NL and in the IL of these rats, the 5-HT concentrations were significantly increased. This indicates that the 5-HT system in the AL reacts differently from those in the NL and IL. Pituitary dopamine (DA) contents were not affected by ether and exsanguination. Thus, the pituitary DA and the pituitary 5-HT neuronal systems are activated independently in different endocrine states. The concentrations of 5-HT were also measured in 10 groups of normal male or female rats in the course of 2 years and compared with those of DA. The concentrations of 5-HT were lowest in the AL with little variation between the different groups. In the rest of the tubero-hypophyseal system the concentrations of 5-HT were at least 3, and up to 10 times, higher than in the AL, with larger inter group variations. The DA concentrations in the AL were only 10-20% of those of 5-HT; in the other regions they were equal to or higher than those of 5-HT.

Immunoreactive substance P in the tubero-hypophyseal system of the rat: selective decrease in the neural lobe after dehydration and sodium loading.

In the tubero-hypophyseal system of the rat the approximate concentrations of immunoreactive substance P (I-SP), expressed as fmol/mg tissue, were 100 in the medial basal hypothalamus (MBH), 20 in the anterior lobe (AL), 20 in the neural lobe (NL) and 4 in the intermediate lobe of the hypophysis. These values were not altered by treatment with capsaicin on day 2 after birth. In rats in which the secretion of neurohormones from the NL was increased by dehydration followed by sodium loading there was a fall by 70% in the I-SP concentration of the NL; no change occurred in the AL or the MBH.

Regional differences in the effect of pargyline on dopamine concentrations in the rat hypophysis.

Dopamine concentrations were estimated in different regions of the infundibular-pituitary system of the rat. The highest values were found in the pituitary stalk where they are comparable to the concentrations in the striatum, and higher than in the median eminence. In the anterior lobe the dopamine (DA) concentrations of the upper pole (10% of the lobe tissue) were 2-5 times higher than in the rest of the lobe. Anterior lobe DA was significantly higher in rats about 6 months old than in rats about 3 months old. After inhibition of MAO, the DA concentrations in the isolated neural lobe (NL) were more than doubled. In contrast there was no increase in the DA concentrations of the intermediate lobe, an observation which is in agreement with our previous finding that DA synthesis in the intermediate lobe after electrical stimulation of the pituitary stalk is much slower than in the neural lobe. Pargyline treatment caused an increase in the DA concentrations in the anterior lobe by about 150%. This rise (together with a DA increase in the stalk-infundibulum) may be related to the decrease in circulating prolactin which occurs in pargyline treated rats.

Dopamine release and synthesis in the neurointermediate lobe of the rat hypophysis in vitro after electrical stimulation of the pituitary stalk.

The isolated neurointermediate lobe of the rat hypophysis was incubated in Krebs solution and the stalk was electrically stimulated. The endogenous dopamine (DA) released into the medium was estimated by HPLC with electrochemical detection. Stimulation with biphasic pulses (1 ms, 10 Hz) in the presence of pargyline elicited a mean DA output of 200 fg X pulse-1. This release was calcium-dependent but was only partially inhibited by tetrodotoxin (TTX) (1 microM), effects typical for direct electrical depolarization of the nerve endings. Reducing the duration of the electric pulses to 0.2 ms (15 Hz) caused a reduction in DA output to about 40 fg X pulse-1 which was completely blocked by TTX indicating that it was evoked by propagated action potentials. DA overflow was enhanced when the action potential was prolonged with tetraethylammonium (TEA) or when DA uptake was inhibited with GBR 12921. Evidence for a calcium-dependent increase in DA synthesis in electrically stimulated NILs has been obtained when monoamine oxidase was inhibited and TEA or GBR 12921 was present in the Krebs solution. The present results complete the requirements for the DA in the NIL to be classified as a neurotransmitter substance. The NIL-pituitary stalk preparation is a useful model for studying regulatory mechanisms in dopaminergic nerve terminals.

Specific changes in type A and B monoamine oxidase activity in different tissues of hypophysectomized rats.

MAO activity was tested in four organs of young male rats 16 days after hypophysectomy. Five monoamines were used as substrates. In the adrenal glands and the thyroid gland MAO activity towards the substrates preferably deaminated by MAO-A was severely decreased. In the adrenal glands, but not in the thyroid gland, the effect of hypophysectomy could be simulated with dexamethasone, a steroid which inhibits ACTH secretion. A significant rise in MAO activity towards substrates for MAO-A and MAO-B occurred in the heart of hypophysectomized rats. There was little change in the MAO activity of whole brain homogenates after hypophysectomy.

A comparison of the release of endogenous dopamine from the neural and the intermediate lobe of the rat hypophysis after electrical stimulation of the stalk.

Dopamine (DA) was released from the isolated neural lobe of the pituitary gland by electrical stimulation of the pituitary stalk. This observation completes the evidence for DA to be classified as a neurotransmitter substance in the neural lobe in the rat hypophysis. From the experiments on the isolated neural lobe and simultaneous experiments on the combined neuro-intermediate lobe it could be calculated that the DA release from the intermediate lobe was about three times larger than that from the neural lobe. Electrical stimulation of the pituitary stalk in the presence of pargyline and TEA led to an increase in DA synthesis in the neural lobe. This effect did not occur in the intermediate lobe.

In vitro release of endogenous catecholamines from the neural and intermediate lobe of the hypophysis.

This paper shows for the first time that the dopamine present in the neural and intermediate lobe (NIL) of the pituitary gland can be released in vitro by depolarization of the tissue with increased [K+]. The release is Ca++ dependent. This observation is in accordance with the concept that dopamine has a function as a neural transmitter substance in the NIL. At a [Ca++] of 2.5 mM it was also possible to measure the norepinephrine release from the NIL. The dopamine release from the NIL was unaffected by GABA. In contrast, the release of dopamine (but not of norepinephrine) from the medial-basal hypothalamus was enhanced by GABA.

Observations on pituitary and cerebral catecholamines in Brattleboro rats.

The combined neural and intermediate lobes (N.I.L.) of the pituitary glands of Brattleboro rats contained significantly more noradrenaline (NA) than those of normal controls of the same age. In addition, in 4 and 6 months old rats the N.I.L. of the homozygotes (Di-rats) contained about twice as much NA as those of the heterozygotes (non-Di-rats). In the enlarged and hyperaemic neural lobes of the Di-rats enlarged blood vessels could be seen. They were surrounded by a rich network of nerve fibres which displayed catecholamine fluorescence. The NA concentrations in the medial basal hypothalamus (M.B.H.) of 4, 6, and 9 months old Di-rats were significantly higher than those of the non-Di-rats and of controls of the same age. This may contribute to the diminished ACTH secretion in Brattleboro rats. The total amount of DA contained in the N.I.L. of 9 months old Di-rats was 20% lower than in the non-Di-rats of the same age. The concentrations of homovanillic acid (HVA) in the corpora striata of 4 months old Di- and non-Di-rats was significantly higher than in older Brattleboro rats and in control rats of various strains and ages. At 9 months it was nearly twice as high in the Di-rats as in the non-Di-rats. The concentrations of dihydroxyphenylacetic acid in the striata of 4--6 months old Di- and non-Di-rats were higher than those of Long Evans rats of the same age.