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The purposes of this study were to assess intra- and interrater reliabilities by novice users of the Myotonometer (Neurogenic Technologies, Inc., Missoula, MT), a portable electronic device that quantifies muscle tone (stiffness) and paresis, in assessing children with cerebral palsy. Two raters used the Myotonometer to assess the biceps brachii and medial gastrocnemius muscles of 10 children with spastic-type cerebral palsy. Muscles were measured in a relaxed state and during a voluntary isometric contraction. Intraclass correlation coefficients and repeatability coefficients were calculated for each muscle and for each condition (relaxed and contracted). Intrarater reliabilities ranged from 0.82 to 0.99 (biceps brachii muscles) and 0.88 to 0.99 (medial gastrocnemius muscles). Interrater reliabilities ranged from 0.74 to 0.99 (biceps brachii muscles) and 0.84 to 0.99 (medial gastrocnemius muscles). Repeatability coefficients indicated a 98% level of agreement between raters across all conditions. Novice users of the Myotonometer, with few exceptions, had high to very high intra- and interrater reliabilities for measurements of the biceps brachii and medial gastrocnemius muscles of children with spastic-type cerebral palsy.

Objectives: To assess the intra- and interrater reliabilities of the Myotonometer, a hand-held, computerized, electronic device that quantifies muscle stiffness (tone/compliance)., Design: Reliability study., Setting: Research laboratory., Participants: Thirty-five healthy, nondisabled adults (age range, 22-42 y)., Interventions: Not applicable., Main Outcome Measures: Two raters used the Myotonometer to evaluate subjects' lateral gastrocnemius and biceps brachii muscles. Muscles were measured in a relaxed state and during a voluntary isometric contraction. Coefficients were calculated for each muscle and each condition (relaxed, contracted). Results were analyzed by using Design II intraclass correlation coefficients., Results: Reliability coefficients were highest when the instrument exerted moderate to strong forces against the muscle (range, 0.50-2.00 kg; intrarater reliability R range, .84 - .99; interrater reliability R range, .75 - .96)., Conclusions: Myotonometer measurements had high to very high intra- and interrater reliabilities for measurements of the lateral gastrocnemius and biceps brachii muscles.

In the turkey, the onset of incubation behavior is associated with altered ovarian steroidogenesis, ovarian regression, decreased, LH secretion, and increased serum prolactin (Prl) levels. To clarify the relative contribution of circulating LH and Prl to the initiation of ovarian regression, laying hens were exposed for 0, 3, 7, or 14 days to a forced molting procedure (exposure to reduced day length of 6L:18D and removal of feed and water for the initial 3 days) that induces ovarian regression and decreased LH levels but does not increase Prl levels. On each of these days, hens were killed and granulosa and theca interna cells from the largest (F1) and fifth largest (F5) preovulatory follicles and total cells from the small white follicles (SWF) were incubated for 5 h in the presence or absence of ovine LH (oLH; 0-1,000 ng/ml). Force-molted hens exhibited diminished levels of circulating LH, Prl, progesterone (P), androgen (A), and estradiol (E) by Day 3 of treatment. Ovarian atresia began in F1 by the third day of treatment, and included F1 and F5 by the seventh day. No preovulatory follicles were present on the fourteenth day. With both F1 and F5 granulosa cells, production of P in the presence of oLH was initially enhanced (Day 3) and later absent (Day 7). In contrast, production of A by F5 theca interna cells in the presence of oLH was initially suppressed (Day 3) and then returned to pretreatment levels (Day 7).(ABSTRACT TRUNCATED AT 250 WORDS)

Three experiments were conducted to evaluate the effect of immobilization on plasma prolactin (PRL) levels in the immature turkey. Acute immobilization for 120 min resulted in elevated PRL levels (P less than 0.05) starting 15 min after the onset of immobilization. Release from immobilization caused PRL levels to return to those of the nonimmobilized controls by 120-180 min following replacement of the turkeys in cages. Repeated immobilization for 3 or 5 consecutive days diminished PRL response (P less than 0.05) to subsequent immobilization. It is suggested that the PRL controlling mechanism(s) of the young turkey is both susceptible and able to habituate to changes induced by immobilization stress.

Porcine vasoactive intestinal polypeptide (VIP; 10(-9) to 10(-7) M) was a potent stimulator of prolactin (PRL) release by anterior pituitary cells from immature and laying turkey hens. Basal and VIP-induced PRL release of cells from laying hens were diminished (P less than 0.05) when the cells were cultured for 48 hr in the presence of charcoal-stripped laying hen serum, but not when the cells were cultured in the presence of whole laying hen serum. This change in VIP-induced PRL release was not evident when cells were derived from immature hens. Basal PRL release by cells from laying hens was not altered by the presence of estradiol (E2; 10(-12) to 10(-5) M), although such release was generally enhanced in cultures of cells from immature hens containing E2. The presence of E2 enhanced (P less than 0.05) the magnitude of the VIP-induced PRL release by cultures of cells from laying hens and diminished (P less than 0.05) the magnitude of this release in cultures of cells from immature hens. Cells from immature and laying hens exposed to progesterone (P4; 10(-5) M) for 96 hr exhibited enhanced basal PRL release, though lower P4 concentrations had no effect. Utilizing cells from laying hens, P4 exposure for 24 hr resulted in diminished (P less than 0.05) VIP-induced PRL release, while P4 exposure for 96 hr resulted in markedly enhanced (P less than 0.05) VIP-induced PRL release. Basal PRL release was generally not altered by the presence of testosterone (T). The VIP-induced PRL release by cells derived from immature and laying hens was diminished (P less than 0.05) by the presence of T. Prolactin release in the turkey is likely modulated by gonadal steroids acting directly on the cells of the anterior pituitary.

Intravenous administration of 1.0 or 3.0 eq hypothalamic extract (HE) to 8-week-old male and 7-week-old female turkeys, respectively, induced an increase in circulating prolactin (PRL) levels but had no effect on circulating luteinizing hormone (LH) levels. The incubation of dissociated anterior pituitary cells from 13-week-old female turkeys with HE induced a dose-related increase in PRL release; however, only the highest dose of HE induced an increase in LH release. Coincubation of a hypothalamic fragment with anterior pituitary cells from 9-week-old females induced a release of both PRL and LH. Dissociated pituitary cells from 11-week-old females initially incubated for 3 hr in medium containing charcoal-treated (stripped) turkey serum yielded a larger release of PRL and LH in the presence of HE than did cells initially incubated with turkey serum or no serum. Luteinizing hormone-releasing hormone (LHRH) induction of LH release was greatest from cells initially incubated with stripped serum for 3 or 24 hr. The LHRH-induced LH release was completely blocked in cells initially incubated for 24 hr with turkey serum. The initial incubation of cells for 24 hr with stripped serum yielded a larger release of PRL and LH in response to HE than did cells initially incubated with serum. The hypothalamus of the young turkey contains substantial PRL-releasing activity as well as LH-releasing activity. The ability of the releasing factors to stimulate pituitary hormone release is influenced by factors present in the blood of the young turkey. This is especially evident in the LHRH-induced LH release where serum factors inhibited the release.

Cytosolic progesterone receptors (PRcs) from the reproductive tract of the female turkey were analyzed by high-performance liquid chromatography using a diethylaminoethyl (DEAE) ion-exchange column. PRcs from oviduct tissue of laying, incubating, photorefractory and short-day turkey hens were compared. In general, three types of PRcs were identified: Receptor I, a partially displaceable species that was eluted at a 0.13 M salt concentration; and Receptors II and III, which were two specific binding species eluting at 0.23 M and 0.26 M, respectively. In the subdivided tissue from the laying hen oviduct, Receptor I was the major PRc species of the isthmus and Receptor III was the only receptor present in the uterus. The infundibulum and magnum each contained a small amount of Receptor II and a substantial amount of Receptor III. The whole oviduct of incubating hens contained a greater proportion of Receptor I than Receptor II or III, and these last two receptor types were present in equal quantity. The whole oviduct of the short-day hens had an equal distribution of the three receptor types. In the presence of sodium molybdate, an inhibitor of phosphatase and protease, only one sharp Receptor II species was seen in the magnum and uterus of the laying hen oviduct and in the whole oviducts of incubating and short-day hens. The transformation of Receptor II to Receptor III in the absence of sodium molybdate was facilitated by the aging of cytosol at 0-4 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

An ambient temperature of 30 degrees C compared to 18 degrees C accelerated the increase in serum prolactin (Prl) level induced by photostimulation of female turkeys. The contribution of reproductive stage and nesting behavior to this serum Prl elevation was assessed by housing adult female turkeys in individual wire cages while allowing other females free access to nests on the floor. Birds of both groups were exposed to 10 degrees C, 24 degrees C or 30 degrees C beginning 4 wk prior to photostimulation and continuing throughout the reproductive phase. Lapsed time between the onset of photostimulation and onset of sexual maturity, and between the onset of sexual maturity and onset of incubation behavior was shorter in birds housed at 30 degrees C with access to nests than in corresponding birds housed at 24 degrees C and 10 degrees C. The increases in serum Prl associated with sexual maturity or incubation behavior occurred at a greater rate in the birds maintained at 30 degrees C. Cage-reared birds had the same lapsed time between onset of photostimulation and onset of sexual maturity and the same sustained low Prl level regardless of ambient temperature exposure. All groups exhibited similar luteinizing hormone profiles. These findings indicate that the accelerated increase in Prl under elevated temperature in floor-reared turkeys is related to accelerated development of reproductive function, and not the direct effect of ambient temperature on mechanisms controlling Prl.

Cyproheptadine (CHD) at doses of .1 to 10.0 mg/kg body weight produced a dose-related elevation in prolactin (PRL) and no change in luteinizing hormone (LH) levels. Cyproheptadine pretreatment failed to depress the quipazine (serotonin agonist)-induced PRL rise. Rather than inhibiting the serotonergic induction of PRL release in the young turkey, CHD exerted a stimulatory influence on PRL release through another mechanism.

Intravenous administration of 0.025, 0.25, or 2.5 micrograms/kg thyrotropin releasing hormone (TRH) to 4-week-old female turkeys induced a dose-dependent increase (P = 0.004) in serum prolactin (PRL) 15 min post-treatment. Dispersed anterior pituitary cell cultures were utilized to determine the effect of TRH on cellular release of PRL, luteinizing hormone (LH), and growth hormone (GH). In the first experiment, cells from 13-week-old male turkeys were initially incubated for 24 hr in Medium 199 (M-199) plus 10% turkey serum and then placed in M-199 plus 10(-10) to 10(-4) M TRH for 5 hr. Incubation with TRH produced no change in PRL release from that of spontaneous release (P = 0.854). However, 10(-5) and 10(-4) M TRH induced LH release (P less than 0.0001). The TRH-induced GH response was parabolic (P less than 0.0001), with the maximal release at 10(-8) M. The second experiment, utilizing pituitary cells from 7-week-old females, studied these responses on 3, 5, and 7 days of monolayer incubation. TRH failed to induce a PRL release in all tests (P greater than 0.162), although hypothalamic extract induced a large release (P less than 0.0001) of PRL each time. Both 10(-6) and 10(-4) M TRH induced a LH release on Day 3 while only 10(-4) M did so on Day 5, and none of the doses elicited a release on Day 7. The parabolic GH response generally persisted in all tests.(ABSTRACT TRUNCATED AT 250 WORDS)

The wet weight of the anterior pituitary gland of the domestic turkey increases as the hen progresses through the reproductive cycle. The greatest wet weight was observed with anterior pituitary glands from incubating hens, whose weight was twice that of anterior pituitary glands from nonphotostimulated hens. Anterior pituitary glands from hens in each of the various stages of the reproductive cycle were dissociated and cultured, and subsequently basal and hypothalamic extract (HE)-induced releases of prolactin (PRL) and luteinizing hormone (LH) were compared between cultures containing a defined number of anterior pituitary cells. Basal and HE-induced releases of PRL were greatest (P less than 0.05) in the cultures of anterior pituitary cells derived from incubating hens, with those of laying, photorefractory, and nonphotostimulated hens being successively less (P less than 0.05), respectively. HE-induced release of LH was greatest (P less than 0.05) in cell cultures derived from nonphotostimulated hens, with cultures of cells derived from laying, photorefractory, and incubating hens exhibiting successively smaller releases of LH (P less than 0.05), respectively. The concentration of HE that induced the first significant increase in the release of PRL or LH above that of basal levels also varied with the reproductive stage of the donor hens. The changes in circulating PRL levels during the various stages of the reproductive cycle reflect changes in anterior pituitary sensitivity to hypothalamic releasing activity and/or changes in the PRL releasing capacity of the anterior pituitary. In contrast, there does not appear to be a strong correlation between anterior pituitary LH releasing capacity in vitro and circulating LH levels in the domestic turkey hen.

Two experiments were conducted to evaluate the effect of estradiol benzoate (EB), progesterone (P), ovine prolactin (oPrl), or their combinations on temporal patterns of serum luteinizing hormone (LH) and Prl and on nesting behavior in adult ovariectomized female turkeys. Levels of serum LH were initially reduced (p greater than 0.05) by the steroid treatments, while continuation of treatments induced surges of LH to levels comparable to pretreatment levels. Administration of steroid increased (p less than 0.05) levels of serum Prl, which persisted until termination of treatments. Administration of oPrl had no effect on levels of serum LH but blunted the steroid-induced release of Prl. Neither EB, P, nor oPrl treatments alone nor a combination of EB + P elicited nest occupation. Nest occupation was observed after administration of P only in turkeys pretreated with EB. Administration of oPrl maintained and advanced the P-induced nesting to persistent nesting behavior (incubation behavior). Once persistent nesting behavior was established, hormonal treatments were terminated, yet nesting behavior was maintained and serum samples showed increasing levels of Prl and decreasing levels of LH. It is suggested that incubation behavior in the female turkey is facilitated by the combined action of estradiol, P, and Prl.

To determine what role pituitary responsiveness plays in the suppression of gonadotropin level during incubation in the turkey, the ability of the pituitary to release luteinizing hormone (LH) in response to luteinizing hormone-releasing hormone (LHRH) was compared in incubating, laying, and photorefractory birds. In all three groups, the i.m. injection of LHRH (4 micrograms/kg) increased serum LH levels; however, the LH response was markedly enhanced in the incubating turkeys as compared with the laying (6.6-fold increase over preinjection levels vs. 1.9-fold; p less than 0.05) or the photorefractory birds (9.7-fold vs. 3.1-fold; p less than 0.05). The LHRH-induced LH release was also determined in turkeys as they shifted from the laying to the incubating phase of the reproductive cycle. This response increased (p less than 0.05) in magnitude as the birds started to incubate. The high prolactin level of incubating turkeys does not have a depressing effect on LHRH-stimulated LH release; thus, impaired LH response to LHRH is not a mechanism involved in the diminished gonadotropin secretion of incubating turkeys.

There are major changes in circulating luteinizing hormone (LH), prolactin (PRL), estrogens (E), and progesterone (P) in relation to the onset of reproduction, egg laying, incubation, and care of young. LH levels increase in the prelaying period, followed some days later by increased circulating levels of E, P, and PRL. Levels of these hormones tend to stabilize during egg laying with periodic ovulatory cycle changes. Around the onset of incubation PRL levels increase, while LH, E, and P levels fall. During incubation PRL reaches very high levels, falling sharply when incubation is terminated. Stimulatory effects of hypothalamic neurotransmitters, peptides, and ovarian steroids on PRL secretion have been shown. The prelaying increase is dependent on E and P and the high levels of incubation require a functional serotonergic system. The causal relationships and roles of PRL in incubation of gallinaceous birds are, however, still unclear.

Serum prolactin (PRL) levels were determined by homologous radioimmunoassay in 6- to 10-week-old domestic white turkeys treated by intraperitoneal injection of agents that alter serotonergic activity. Quipazine (0.1-10.0 mg/kg), a serotonin (5-hydroxytryptamine; 5-HT) agonist, induced a dose-dependent rise in serum PRL level 1 hr after injection. The 5-HT precursor, 5-hydroxytryptophan (5-HTP), at doses of 80 and 150 mg/kg produced over a twofold elevation in PRL level 1 hr after administration, though the 50 mg/kg dose failed to produce any change. Administration of fluoxetine (10 mg/kg), a 5-HT reuptake blocker, induced an elevation in PRL level persisting 3 hr. When fluoxetine injection preceded administration of a weakly stimulatory dose of 5-HTP, a prolonged elevation in PRL level was observed. methysergide (MES), a 5-HT antagonist, administered in a 10 mg/kg dose produced no change in PRL level, while the 25 mg/kg dose initially produced a spike in PRL level which subsequently dropped slightly below the control level. Prior injection of 20 mg/kg MES completely blocked the serum PRL rises induced by quipazine and 5-HTP. These results suggest that serotonergic mechanisms are involved in the regulation of pituitary PRL release beyond basal levels in young domestic turkeys.

Nest deprivation of incubating turkeys caused a decrease in serum prolactin (Prl) levels from 1184.5 +/- 116.4 ng/ml to 896.8 +/- 83.0 ng/ml 1 day after initiation of deprivation, with a further decline to 156.5 +/- 111.7 ng/ml at the end of the 22-day experimental period. Serum luteinizing hormone (LH), progesterone and estradiol levels following nest deprivation were similar to those in birds allowed to incubate (controls). Oral administration of p-chlorophenylalanine (PCPA, 50 mg/kg) to incubating turkeys for 3 consecutive days reduced nesting frequency (P less than 0.05) on the 4th day after initiation of treatment and the nesting virtually ceased by the 9th day. Pretreatment Prl was 1655 +/- 210 ng/ml and declined (P less than 0.05) after PCPA administration to a low of 28.6 +/- 2.8 ng/ml. In addition, PCPA caused a sustained rise in serum LH peaking (5.59 +/- 1.09 ng/ml) 3 days after treatment initiation. Contrary to nest deprivation, serum levels of progesterone and estradiol increased (P less than 0.05) as a consequence of PCPA treatment. Seven of 8 PCPA-treated birds later came into lay when their Prl levels and nesting frequency increased again. The results suggest a role for serotonin (5-HT) in incubation behavior, and Prl and LH secretion in turkeys.

The involvement of catecholamines (CAs) in the control of prolactin (PRL) secretion was investigated in female turkeys. The birds were treated with DL-alpha-methyltyrosine methyl ester HCl (alpha-MT; 10, 100, or 200 mg/kg), diethyldithiocarbamate (DDC; 200 or 400 mg/kg), or phentolamine (5, 10, and 20 mg/kg), all antiadrenergic drugs. Reduction in central CAs by alpha-MT or DDC, or blockade of adrenergic receptors by phentolamine resulted in elevated serum PRL levels in a dose-dependent manner. Prior administration of the adrenergic stimulant clonidine (15, 150, or 1500 micrograms/kg) attenuated the elevated PRL response to phentolamine (20 mg/kg). These results are consistent with the view that the release of PRL in the turkey may be under the inhibitory influence of an adrenergic system.

Serum growth hormone (GH) and luteinizing hormone (LH) levels were determined by homologous radioimmunoassay in 8 to 10-week-old domestic white turkeys treated by intraperitoneal injection of agents that alter serotonergic activity. The serotonin, 5-hydroxytryptamine (5-HT) precursor, 5-hydroxytryptophan (5-HTP), at doses of 80 and 150 mg/kg, produced a dose-related elevation in serum GH (P = .09) and no change in serum LH (P = .30). Administration of fluoxetine (10 mg/kg), a 5-HT reuptake blocker, produced no change in serum GH or LH. When fluoxetine injection preceded the administration of a nonstimulatory dose of 5-HTP, an elevation in GH was observed, but again no change in LH level was evident. Stimulation of postsynaptic 5-HT receptors with quipazine (.5 to 10.0 mg/kg) resulted in a similar elevation in GH; persistence of the effect was dose dependent. Quipazine administration had no influence on serum LH levels. Administration of methysergide (10 or 25 mg/kg), a 5-HT antagonist, induced no change in either serum GH or LH levels. Serotonergic input appears to stimulate GH release and has no apparent influence on LH release in young turkeys.

Serum prolactin (PRL) and luteinizing hormone (LH) levels from mature female turkeys were determined following ovariectomy and daily subcutaneous injections for 4 or 9 days of 0.002-2.0 mg/kg estradiol benzoate (EB), 0.05-2.0 mg/kg progesterone (P), or their combinations. Serum PRL levels were increased (P less than 0.05) at the onset of sexual maturity in intact controls, but not in ovariectomized turkeys. Injection of 0.02 mg/kg EB into ovariectomized turkeys resulted in elevated (P less than 0.05) serum PRL levels after 8 treatment days. Higher doses failed to elicit a response of greater magnitude. EB at doses of 0.02-2.0 mg/kg reduced LH levels while 0.002 mg/kg EB increased LH levels above (P less than 0.05) those of ovariectomized controls. The 1.0-mg/kg P dose increased (P less than 0.05) serum PRL within 24-48 hr of administration while the 0.05-, 0.5-, or 2.0-mg/kg P doses failed to elicit a response. When P was injected in doses of 0.5-2.0 mg/kg, LH levels were decreased in a dose-response manner. The injection of EB combined with P had a variable effect on serum PRL levels. The 0.5- or 1.0-mg/kg dose of P facilitated (advanced in time) the rise in serum PRL level induced by 0.02 mg/kg of EB. In contrast, the positive feedback effect of 0.2 mg/kg of EB was blocked when administered with 0.5 mg/kg of P. Serum LH levels were dramatically decreased by all steroid combinations used. These results indicate that daily injections of EB and/or P in ovariectomized turkeys have a variable effect on serum PRL and LH levels depending upon the dose, the duration of treatment, or the ratio between EB and P.

The magnitude of luteinizing hormone (LH) release during a 3-hr test incubation was diminished (P less than 0.05) when anterior pituitary cells from young turkeys were cultured for 24 to 120 hr. This trend was evident with basal LH release and with LH release induced by luteinizing hormone-releasing hormone (LH-RH) or hypothalamic extract. Anterior pituitary cells were cultured with various concentrations (10(-14) to 10(-6) M) of estradiol (E2), progesterone (P4), or testosterone (T) for 24 hr and then exposed to LH-RH or control medium for 3 hr, still in the presence of steroids. Basal LH release was potentiated (P less than 0.05) when cells were cultured with 10(-8) or 10(-6) M T, but not with E2 or P4. When cells were cultured with E2, LH release in the presence of 10(-8) M LH-RH was enhanced (P less than 0.05) in a dose-dependent fashion. LH-RH mediated LH release was also enhanced (P less than 0.05) when cells were cultured with 10(-8) M P4 or 10(-6) M T. Gonadal steroids can act directly on the anterior pituitary of the young domestic turkey to modulate LH release, with T enhancing basal LH release and E2 potentiating LH-RH-mediated LH release.

Immature female turkeys (7 wk old) were treated with 5-azacytidine, a cytidine analog that has been shown to inhibit cytosine methylation in eukaryotic DNA. Such methylation has been repeatedly implicated in the control of specific gene expression in higher vertebrates. Turkeys administered 5-azacytidine (5 mg/kg) by intravenous injection daily for 5 days exhibited elevated (P less than .05) circulating prolactin (Prl) levels, whereas hens treated with saline, cytidine, or 6-azathymine did not show altered circulating Prl levels. Absence of response upon treatment with the normal nucleoside cytidine or the 6-aza analog of thymine reasonably eliminates nonspecific drug-induced stress as the cause of Prl level elevation. Based on these findings, DNA methylation is implicated in the control of circulating Prl levels in the young turkey.

Primary anterior pituitary cell cultures were utilized to study the influence of serotonin (5-HT) directly on the pituitary. Cells incubated with 10(-5) and 10(-4) M 5-HT exhibited a significant prolactin (Prl) release, whereas cells incubated with 10(-10) to 10(-6) M 5-HT did not. Cells incubated with 10(-10) to 10(-4) M quipazine (5-HT agonist) or methysergide (MES; 5-HT antagonist) did not release Prl in amounts greater/less (P greater than 0.01) than spontaneous release. Luteinizing hormone (LH) release from cells incubated in the presence of 5-HT, quipazine, or MES was similar to spontaneous release. The hypothalamic extract-induced Prl and LH release from cells was not influenced by quipazine, but Prl release was diminished in a dose-related fashion by MES. The influence of 5-HT on hypothalamic induction of Prl and LH release was investigated utilizing in vitro culture of hypothalamic fragments (HF). Media samples from HF incubated with 10(-6) and 10(-4) M 5-HT induced a release of Prl. Media samples from HF incubated with 10(-4) M MES induced less Prl release than media samples from control fragments. When HF were incubated with both 10(-4) M 5-HT and 10(-4) M MES, the expected 5-HT-mediated Prl release was not evident. These culturing situations had no influence on LH release. In vitro Prl release from pituitary cells of the young turkey was stimulated through 5-HT activity at the hypothalamus, but not by direct 5-HT action on the pituitary cells.