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Institut National de la Recherche Agronomique, Unité de Recherche Associeé Centre National de la Recherche Scientifique 1291, Laboratoire de Neuroendocrinologie Sexuelle, 37380 Nouzilly, France
Address all correspondence and requests for reprints to: Dr. Benoît Malpaux, Institut National de la Recherche Agronomique, PRMD, Laboratoire de Neuroendocrinologie Sexuelle, 37380 Nouzilly, France. E-mail: malpaux{at}tours.inra.fr
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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-methyl-paratyrosine [MPT, an inhibitor of
tyrosine hydroxylase (TH); 10 mM in Ringer’s lactate] was
infused into the ME for 5 h, preceded by a 5-h control period
during which only vehicle was infused, in each of the three
photoperiodic treatments. MPT dramatically decreased the
3,4-dihydroxyphenylacetic acid concentration, similarly in all three
photoperiodic treatments, suggesting a similar inhibition of TH
activity. In the LD1 treatment, MPT significantly increased LH pulse
frequency (+1.22 ± 0.46 pulse/5 h from control period, mean
± SEM, n = 9; P < 0.05) and mean
concentration (+51 ± 28%; P < 0.001). In
the other two photoperiodic treatments, MPT had no significant
effect on LH release. Thus, blockade of dopamine synthesis in the ME
seems to stimulate LH secretion in early, but not long-term, inhibition
by LD nor after the transition to short days. Therefore, dopaminergic
activity of the ME seems to be critical for LH secretion inhibition in
some photoperiodic inhibitory treatments but not in others. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Dopaminergic terminals, located in the external layer of the median
eminence (ME) (8), establish contacts with LHRH terminals (9),
providing anatomical evidence for a DA role in the regulation of LH
secretion. Furthermore, DA content in the ME is lower under SD than LD
(10) and a 1.5-fold decrease of the tyrosine hydroxylase (TH,
rate-limiting enzyme of catecholamine biosynthesis) activity is
observed when LH secretion increases after a transfer from LD to SD
(11). More recently, it has been shown that a blockade of DA synthesis
in the ME, by local infusion of -methyl-para-tyrosine (MPT, a
competitive TH inhibitor) stimulates LH secretion in LD-inhibited ewes
(12). Taken together, these studies provide strong evidence that the ME
dopaminergic terminals are involved in the photoperiod-induced
inhibition of LH release.
It is important to note that the neuroendocrine mechanisms leading to
inhibition of LH secretion may vary according to the stage of
reproductive inhibition. For example, injection of the DA antagonist,
pimozide, stimulates LH secretion in ewes during the onset of
photorefractoriness to artificial SD, but has little or no effect in
artificial LD or natural photoperiod inhibited ewes (13, 14).
Similarly, injection of pimozide induces a greater increase in LH
secretion in early anestrus, compared with injection later in this
season (15). Also, lesions in the anterior hypothalamic area delay, but
do not suppress, the inhibitory effect of a shift from SD to LD, and
they delay or totally suppress the apparition of photorefractoriness to
SD (16). These experiments suggest that several neuroendocrine
mechanisms may promote the same physiological state, i.e.
inhibited LH secretion, and that the functional relative importance of
each of these mechanisms for LH inhibition may vary according to the
duration of the inhibition and/or the photoperiodic history. The
objective of our study was to test the hypothesis that functional
significance of ME dopaminergic activity varies according to the
duration of the inhibition and the photoperiodic history. Accordingly,
we examined the effect on LH secretion of a local delivery of MPT in
the ME at three different times of the photoperiod-induced LH secretion
inhibition: after a short-term LD inhibition (after 33 LD), where it
was shown that TH activity is critical for LH secretion inhibition
(12); after a longer exposure to LD (after 72 LD); and during the
interval between the start of SD exposure and the beginning of the
stimulation of LH secretion (after 34 SD).
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Experimental design
Infusion of MPT in the ME was performed using reverse
microdialysis in three photoperiodic treatments where LH secretion is
inhibited: 1) 33 days after the shift from SD to LD (LD1 treatment),
i.e. at the very beginning of LD-induced inhibition of LH
secretion [in this treatment, it is known that blockade of TH in the
ME induces a stimulation of LH secretion (12)]; 2) 72 days after the
transition from SD to LD (LD2 treatment), i.e. after a
longer term inhibition by LD; and 3) 34 days after the transition from
LD to SD (SD treatment), i.e. during the interval between
the start of SD exposure and the beginning of stimulation of LH
secretion. For technical reasons, microdialysis sessions were performed
over a period of 2–4 days for each photoperiodic treatment.
The ewes were allocated to three groups, which were maintained under
the same photoperiodic conditions. The animals entered
photoperiod-controlled rooms on January 26 and underwent two cycles
of artificial light regimen, alternating 105 day periods of constant
LD (16 h of light, lights on at 0600 h) and constant SD (8 h of
light, lights on at 0700 h). On Day 0 (end of the second SD
period), the photoperiod was switched to LD, which was maintained for
77 days. It was then switched to SD for 89 days and then to LD again
(Fig. 1
). Each group underwent
microdialysis treatments twice, in two different photoperiodic
treatments, as described in Fig. 1. In each photoperiodic treatment,
half of the animals were dialyzed for the first time and half for the
second time. This protocol was built to limit the number of
microdialysis sessions on each animal while controlling for the
repetition of this treatment.
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MPT. This dose produces a marked
inhibition of LH secretion during anestrus (12), but the suppression is
not complete. The presence of endogenous LHRH and LH pulses during the
control period before MPT challenge is important to maintain
pituitary responsiveness and to obtain a good consistency between LHRH
and LH pulses in response to an acute treatment. In addition, this
experimental model seems to be more physiologically relevant, because
the inhibition of LH secretion obtained with 0.5 cm implant is similar
to the inhibition observed during seasonal anestrus in intact animals
(18). For the experiment, the ewes were ovariectomized at least 6 weeks before the first microdialysis session and, at the same time, implanted with two sc Silastic estradiol implants (0.5 cm and 1.5 cm in length). This dose of estradiol was maintained before and between the microdialysis sessions. The 1.5-cm estradiol implant was removed 72 h before microdialysis session, whereas the 0.5-cm implant was left in place. At the end of the session, the 1.5-cm implant was replaced.
Surgical access to the ME
At least one month before microdialysis, animals were implanted
with a stainless steel guide cannula (external diameter, 1.2 mm;
internal diameter, 0.8 mm; length, 55 mm), aimed toward the ME,
according to a surgical procedure described previously (12).
The site targeted was located 1 mm in front of the tip of the infundibular recess and 1 mm below the third ventricle floor. The laterality was 0.8–1.0 mm from the middle of the third ventricle. The tip of the guide cannula was left 6 mm above the site.
At the end of the surgery, a stainless steel stylet, of the length of the cannula, was inserted in it. It was removed to allow placement of the microdialysis probe and was replaced between two dialysis sessions.
Microdialysis sessions
The day before each microdialysis session, animals were placed
in individual pens so that they could see each other. At this time,
they were equipped with a harness to support the microdialysis pump,
and a polyethylene catheter was inserted in the jugular vein.
At least 12 h before the beginning of the blood sampling, the microdialysis probe was inserted in the guide cannula and infused with Ringer’s lactate at a slow rate (0.5 µl/min) overnight. The probes were built according to a procedure described previously (19). The length of the active membrane was 3 mm.
About 1 h before the first blood sample, on the day of the
experiment, the flow rate was increased to 1 µl/min. The experiment
began at 0900 h and lasted 10 h, divided into 5 h of
control period (infusion of Ringer’s lactate) and 5 h of MPT
infusion (10 mM in Ringer’s lactate). The drug was diluted
1 h before use. To assess the effect of MPT on the metabolism
of the different monoamines, dialysates were collected into glass tubes
containing 4 µl of 0.25 M perchloric acid. The tubes were
changed every 30 min and immediately frozen in dry ice and were stored
at -20 C until assay. Blood samples (10 min) were collected into
heparinized tubes and centrifuged, and plasma was removed for later RIA
to assess the effect of MPT on LH secretion. Because PRL secretion
is strongly regulated by photoperiod (20) and DA is involved in this
regulation (21), PRL was also assayed in every third sample.
Histological processing of the brains
Ewes were killed, with their probes left in place, 1 or 2 days
after their last microdialysis treatment. Brains were processed,
according to a procedure described previously, to locate the position
of the microdialysis probes within the tissue (12). Coronal sections
(40-µm thick) of the hypothalamic block containing ME were collected
and stained with cresyl violet acetate, as described previously
(22).
Hormonal and catecholamine assays
LH was assayed in duplicate 100-µl aliquots of plasma using
the RIA of Pelletier et al. (23), modified by Montgomery
et al. (24). Sensitivity (2SD from buffer
controls) was 0.1 ± 0.0 ng/ml (mean ± SEM) of
1051-CY-LH (0.2 ng/ml of NIH-LH-S1). The intraassay CV for three plasma
pools averaged 8.3%, and the interassay CV for these plasma pools
averaged 16.1% (2 assays).
PRL was assayed in duplicate 10-µl aliquots of plasma using the RIA of Kann (25). Sensitivity was 8.5 ng/ml of NIDDK-oPRL. The intraassay CV for three plasma pools averaged 9.5%, and the interassay CV for these plasma pools averaged 21.7% (2 assays).
Catecholamines (i.e. DA, adrenaline, noradrenaline), their metabolites [i.e. 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), 3-methoxy-4-hydroxyphenylethyleneglycol (MHPG)], and 5- hydroxy-indoleacetic acid (5-HIAA, a serotonin metabolite) were assayed directly in dialysates using HPLC in semimicrobore conditions (ULTRACARB Phenomenex 5 ODS 20 column) coupled to electrochemical detection. The mobile phase consisted of KH2PO4 (85 g/liter), sodium octane sulfonate (0.093 g/liter), EDTA (0.08 g/liter), and methanol (5%) in 1 liter distilled water, pH = 4.2. The flow rate was 1.2 ml/min. The limit of detection for each compound was defined as the quantity giving a peak with an amplitude equal to twice the amplitude of the baseline noise (4 pg for DA, 5 pg for DOPAC, 2 pg for noradrenaline, 2 pg for adrenaline, 6 pg for MHPG, 8 pg for HVA, 6 pg for 5-HIAA).
Data analysis
Concentrations of LH, PRL, and amines that were lower than
detection limit were assigned assay sensitivity for data analysis and
plotting. Mean LH, PRL, and amine concentrations during the control
period were compared with a two-factor ANOVA (photoperiodic treatment;
rank of dialysis) using SuperANOVA software (Abacus Concepts, Inc.,
Berkeley, CA). This was done as a preliminary analysis to
determine whether the neuroendocrine state at the beginning of the
pharmacological treatment was influenced by the photoperiodic treatment
(factor photoperiodic treatment: LD1, LD2, SD) and whether animals had
or had not been used for microdialysis before (factor rank of dialysis:
first, second). The effect of MPT infusion on mean LH, PRL, and
amine concentrations was determined by a three-way repeated-measures
ANOVA: microdialysis period (control vs. MPT infusion)
was taken as a within factor, and photoperiodic treatment and rank of
dialysis were taken as between factors.
LH pulse detection was performed by the mean of the Munro algorithm
(26), according to the method of Merriam and Wachter (27) (G
parameters: G1 = 3.78; G2 = 2.20; G3 = 1.60; G4 =
1.24; G5 = 0.93; Baxter parameters: b1 = 0.07997; b2 =
0.03; b3 = 0.003). LH pulse frequency was analyzed in a way
similar to that used for mean LH concentration, but with nonparametric
tests. Two preliminary analyses were performed. First, an effect of
rank of microdialysis on the response to MPT was tested on all
animals taken as a group, or in each photoperiodic treatment, by a
Mann-Whitney U test (StatView SE, Abacus Concepts, Inc.)
bearing on the difference of the number of pulses between control and
MPT infusion period. Second, differences between photoperiodic
treatments in LH pulse frequency during the control period were tested
by a Kruskal-Wallis test. In each treatment, we analyzed the effects of
the infusion of MPT on LH pulse frequency, by a Wilcoxon
nonparametric test. Comparison of the effect of MPT between the
different photoperiodic treatments was performed by comparing the
difference (MPT infusion period minus control period)
with a Kruskal-Wallis test, followed by a Mann-Whitney U
test for two-by-two comparisons.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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). In 5 of these 15 animals, the top of
the active membrane touched the bottom of the arcuate nucleus (nos. 2,
5, 9, 18, and 27). In 3 animals (nos. 3, 6, and 20), diffusion in the
third ventricle cannot be excluded because the trace of the dialysis
probe disrupted the ventricle wall, and for 2 of these ewes (nos. 3 and
20), the trace was located in the middle region of the ME. In contrast,
in 13 ewes, the trace of the probe membrane reached the lateral region
of the ME. No evidence for a relationship between location of the probe
within the ME (laterality or anteroposterior plane) and response to
MPT was found in any treatment.
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LH response to photoperiodic treatments
At the time of MPT treatment, LH concentrations were low in all
groups (Fig. 3). Mean LH concentrations
of the last three samples preceding the removal of the 1.5-cm estradiol
implant were not different among photoperiodic situations.
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MPT infusion on these parameters. Furthermore, these parameters
during the control period did not differ among the three photoperiodic
treatments, suggesting that animals were at the same apparent
physiological state before the MPT infusion started (Figs. 4 and 5).
During the control period (0.5-cm estradiol implants), mean LH
concentration averaged 0.88 ± 0.12 ng/ml, with a mean pulse
frequency of 1.80 ± 0.38 pulse/5 h (n = 30; mean of all
animals across all photoperiodic treatments). The increase in mean LH
concentration after the removal of the 1.5-cm estradiol implant was not
different among treatments. Indeed, the difference between mean LH
concentrations during the control period of the microdialysis sessions
(0.5-cm implant) and before removal of the 1.5-cm estradiol implant
(last three blood samples) was 0.42 ± 0.22 ng/ml in LD1 (n =
9), 0.36 ± 0.16 ng/ml in LD2 (n = 13), and 0.35 ± 0.30
ng/ml (n = 8; P > 0.9).
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MPT on monoamine concentrationMPT infusion caused a major decrease in DOPAC concentration
(63 ± 5%, n = 30, mean of all animals across all
photoperiodic treatments; P < 0.001; Fig. 4
) and this
effect was not different among the three photoperiodic treatments,
because no interaction between MPT infusion and photoperiodic
treatment was revealed by ANOVA (P > 0.4). DOPAC
concentration decreased by 67 ± 11% (n = 9) in the LD1, by
60 ± 8% (n = 13) in the LD2, and by 64 ± 8%
(n = 8) in the SD treatments.
About 86% (for all animals across the three photoperiodic treatments)
of the values of DA concentration during the control period were equal
or below the detection threshold. Nevertheless, as for DOPAC, MPT
infusion caused a significant reduction in DA concentration in the
dialysates (11 ± 3 pg/20 µl during control period
vs. 6 ± 1 pg/20 µl during MPT infusion period,
n = 30, mean of all animals across all photoperiodic treatments;
P < 0.05), independent of the photoperiodic treatment
(interaction MPT infusion x photoperiodic treatment:
P > 0.09).
HVA and 5-HIAA were detected in 9 of the 15 animals (i.e. 18
observations out of 30: 7 out of 9 in the LD1 treatment, 9 out of 13 in
the LD2 treatment, and 2 out of 8 in the SD treatment; Chi-squared
= 5.73, P > 0.05). In these animals, even though DOPAC
concentration was affected by MPT infusion (control: 41 ± 10
vs. MPT: 11 ± 3 pg/20 µl; n = 18;
P < 0.05), mean HVA and 5-HIAA concentrations did not
significantly differ between control and MPT infusion periods (HVA:
control: 1180 ± 447 vs. MPT: 1017 ± 419 pg/20
µl; 5-HIAA: control: 163 ± 54 vs. MPT: 148
± 59 pg/20 µl; n = 18). Moreover, no difference in amine
concentrations was found among the three photoperiodic treatments.
Effect of MPT on LH secretion
The effects of MPT on LH secretion in the different
photoperiodic treatments are illustrated by three individual examples
taken from each group of ewes (Fig. 7).
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MPT infusion (P = 0.01).
However, there was an interaction between MPT infusion and the
photoperiodic treatment (P < 0.05). Increase in mean
LH concentration was only significant in the LD1 treatment
(P < 0.001). Two-by-two comparison revealed that the
effect of MPT in this treatment was significantly different from the
two other photoperiodic treatments (P < 0.05; Fig. 5).
For LH pulse frequency analysis, because no effect of rank of
microdialysis treatment was found, all the data (coming from the first
and second dialysis) were pooled for further analysis. MPT infusion
in the ME stimulated LH pulsatility in the LD1 treatment
(P < 0.05; Fig. 5). In contrast, it produced no
significant effect in the LD2 treatment and in the SD treatment. The
effect of MPT on LH pulse frequency, assessed by the difference
between MPT and control periods, was significantly different between
LD1 and LD2 treatments (+1.22 ± 0.46, n = 9 vs.
-0.08 ± 0.18 pulses/5 h, n = 13; P <
0.01). The SD treatment effect was not significantly different from the
other two (+0.63 ± 0.53 pulses/5 h, n = 8).
For pulse amplitude, no significant effect of the MPT infusion was
found in any of the photoperiodic treatments.
Effect of MPT on PRL secretion
Overall, no effect of MPT infusion was found on PRL release
(Fig. 6). A significant interaction between MPT infusion and
photoperiodic treatment was detected (interaction MPT infusion x
photoperiodic treatment: P < 0.05). This interaction
is explained by the fact that MPT infusion tended to cause an
increase in mean PRL concentration in the SD treatment (+46 ±
23%, n = 8; Fig. 6), whereas PRL concentration tended to decrease
slightly in the other two treatments (LD1: -9 ± 5%, n = 9;
LD2: -8 ± 5%, n = 13). Nevertheless, none of these changes
reached significance.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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MPT in the ME, which reduces
TH activity, in photo-inhibited ewes, stimulates LH secretion after 33
days of LD exposure, whereas it does not produce such an effect during
a longer term inhibition by LD or after 34 days of exposure to SD. This
differential effect of MPT is observed despite a similar inhibition
of DA synthesis. Therefore, dopaminergic activity of the ME seems to be
critical for LH secretion inhibition in some photoperiodic inhibitory
treatments and not in others.
MPT infusion induced a dramatic decrease in the extracellular
concentration of DOPAC, which declined in less than 1 h after the
beginning of the infusion and remained suppressed throughout the
infusion. Concomitantly, a significant decrease of DA concentration was
observed. These findings suggest that local inhibition of TH activity
was effective. For DA, many values (about 86%) during the control
period were equal to or below the detection threshold, so the effect of
MPT on DA release was probably underestimated by our measurements.
Consequently, DOPAC was taken as an index for the effect of MPT on
DA synthesis, for two reasons. First, in the rat striatum, infusion of
MPT induces a concomitant dramatic decrease in both DA and DOPAC
concentrations (28), which is verified in the ewe ME in our experiment.
Second, DOPAC is a direct metabolite of DA, and its concentration in
rat ME extracts reflects the rate of DA synthesis in the terminals of
the tuberoinfundibular dopaminergic neurons (29). Analysis of DOPAC
concentration among the different photoperiodic treatments did not
reveal any difference in this metabolite during the control period and,
most importantly, showed that the treatment produced a similar
reduction in DOPAC concentration, regardless of the photoperiodic
treatment. This suggests that the infusion of MPT induced a similar
inhibition of TH activity in the ME in the three photoperiodic
treatments.
Concerning the other monoamines, 5-HIAA was detected in the dialysates
of 9 of the 15 ewes. In these animals, whereas DOPAC and DA
concentrations decreased during the MPT infusion period, 5-HIAA
concentration was not affected, confirming that MPT had no effect on
serotonin metabolism. Adrenaline, noradrenaline, and MHPG were never
detected. This observation is consistent with the low number of
noradrenergic or adrenergic terminals in the ME of sheep, as
demonstrated by a weak immunoreactivity for dopamine-ß-hydroxylase
(DBH) (8). The arcuate nucleus, however, exhibits a high
immunoreactivity for DBH (8), in contrast to the ME; and in some
animals, the upper part of the dialysis membrane reached the arcuate
nucleus. Although we cannot fully exclude that a small diffusion of
MPT into the arcuate nucleus may have contributed to the observed
physiological effects, the absence of noradrenaline and MHPG in the
dialysates suggests that this contribution, if any, was negligible.
Consistent with this conclusion, the infusion of MPT did not produce
a clear effect on PRL secretion, whereas DA cell bodies located in the
arcuate nucleus are implicated in the control of PRL secretion (21).
Also, it can be noted that when MPT infusion stimulated LH
secretion, the stimulatory effect was found only on pulse frequency and
not on pulse amplitude, suggesting that MPT did not have any effect
at the level of the pituitary. Therefore, the blockade of TH activity
seems to be limited to the ME for its major part.
The stimulatory effect of MPT on LH secretion in the LD1 treatment
concurs with previous results (12). Interestingly, our results extend
those of the latter study by showing that the same treatment with
MPT loses its stimulating influence on LH secretion during a longer
term inhibition by LD, or during the lag time before LH is stimulated
after the shift to SD. This contrast was found despite the fact that,
in the three photoperiodic treatments, LH secretion was similar during
the control period, and TH activity was similarly inhibited by the
MPT infusion. These results support the hypothesis that
neurotransmitters different from DA are also involved in transducing
the inhibitory effect of photoperiod on LH secretion, as suggested by
several experiments (13, 14, 15, 16). For example, blockade of DA action by
injection of pimozide produces a small stimulatory effect on LH
secretion in ewes inhibited by a 40-LD exposure, but such a stimulatory
effect is no longer observed in ewes exposed to 60 or 80 LD (13, 14).
Similarly, lesion of the anterior hypothalamic area delays the onset of
inhibition of LH secretion after a transfer from SD to LD but does not
prevent the occurrence of the inhibition, which occurs about 6 weeks
later (16). Moreover, our study extends the previous conclusions by
identifying an anatomical site, ME, where DA contribution for
inhibition of LH secretion varies during the inhibited LH period.
Several hypotheses can be considered to explain the results. The
differential effect of MPT on mean LH levels could reflect a change
in pituitary responsiveness between photoperiodic treatments. Although
we cannot fully exclude this possibility, it seems highly unlikely.
Indeed, the differential effect observed on mean levels is correlated
to and, most likely, the consequence of a change in LH pulse frequency,
which was shown to reflect LHRH pulse frequency in similar situations
of photoperiodic inhibition (1). Furthermore, the low dose of estradiol
used during the 3 days preceding the study maintained a pulse frequency
that was low but sufficient to maintain pituitary responsiveness and
LH-releasable storage (30). Furthermore, pituitary responsiveness is
influenced mainly by two factors (31): the steroid milieu, which was
identical among the three photoperiodic treatments; and LHRH pulse
frequency, which was not different among the situations at the time of
MPT challenge, as assessed through LH pulse frequency during the
control period. Along the same lines, the differential effect of MPT
could be caused by a differential sensitivity to reduction in estradiol
feedback after the reduction of the estradiol treatment, 3 days before
the microdialysis session. This reduced dose of estradiol caused an
increase in LH secretion: mean levels increased from 0.5 ng/ml before
removal of the implant to 0.9 ng/ml during the control period of the
microdialysis session. However, this increase was not different among
treatments, which is not consistent with a differential sensitivity to
the changing estradiol levels. In addition, in the first situation
(LD1), where (according to this hypothesis) a faster reduction in
estradiol feedback would have enhanced the effect of MPT, the
increase in LH pulsatile secretion after MPT treatment is not
related to the dose of estradiol. Indeed, a similar stimulatory effect
of MPT is observed whether animals are treated with 0.5-cm or 2-cm
estradiol capsules in this situation (F. Bertrand, B. Malpaux,
unpublished data).
Therefore, two main hypotheses may explain the differences in the
effects of MPT. The first deals with differences in the TH activity
of the ME. Stimulation of LH secretion in the LD1 treatment could be
caused by a lower TH activity in the ME than in the LD2 and SD
treatments. Thus, the blockade of TH activity by MPT, even partial,
would be sufficient in the LD1 treatment to partly remove
inhibition of LH secretion but not sufficient in the other two
treatments. Nevertheless, direct ME TH activity measurements have shown
that such activity is high in the LD2 and SD treatments (11, 32), and
the similar concentrations of DOPAC in the dialysates during the
control period in the three photoperiodic treatments suggest that it is
also elevated in the LD1 treatment, which makes the hypothesis
unlikely.
The other main hypothesis is that the photoperiod- induced
inhibition of LH secretion results from interactions between different
neuronal systems, and that the interactions between DA and the other
neurotransmitters vary in the different photoperiodic stages. For
example, the absence of stimulated LH release by MPT in the LD2 and
SD treatment could be caused by the action of non-DA inhibitory
systems, which replace or reinforce the inhibitory action of DA. Also,
it could be caused by the absence, in these photoperiodic treatments,
of neuronal inputs necessary for a stimulation of LH secretion. In
favor of the hypothesis that photoperiod-induced changes in LH
secretion may also involve neurotransmitters other than DA, evidence
for the implication of serotonin (33, 34), noradrenaline (35), and
excitatory amino-acids (36, 37, 38) has already been obtained. Also, the
number of neuropeptide Y immunoreactive cells is higher in SD than in
LD, and interestingly, in relation to our study, this change takes
place in the ME (39). Some authors postulate that inhibition of LH
secretion in natural anestrus could be maintained by a sequential
process including two components: photorefractoriness (to SD) and
photosuppression (by LD) (40), and that these two components could be
supported by different neuroendocrine mechanisms (14). Injection of
pimozide stimulates LH secretion in a similar manner in ewes refractory
to SD (13) and during the early anestrus season (14) and, most
importantly, has no more effect later in the anestrus season (15),
consistent with the hypothesis that initiation of anestrus involves a
DA-dependent photorefractory phase. Extending this reasoning, we could
postulate that inhibition by LD follows the same scheme, with an early
phase dependent on the DA in the ME and responsible for initiation of
LH secretion inhibition, and a later phase contributing for the
maintenance of LH secretion inhibition and for which ME DA is not the
main inhibitory system.
In conclusion, we have shown that the DA activity of a specific anatomical structure, the ME, does not always transduce the inhibitory effect of photoperiod on LH secretion. These results provide new insights in the field of neuroendocrine regulation: an apparently identical physiological state, photoperiod-inhibited LH secretion, may result from changing interactions between several neuroendocrine systems, of which individual contributions vary during the inhibitory period.
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Acknowledgments |
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Footnotes |
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2 Supported by a Ph.D. grant from Ministère de l’Education
Nationale, de la Recherche et de la Technologie.
3 Present address: Reproductive Sciences Program, 11th Floor, 300
North Ingalls Building, Ann Arbor, Michigan 48109-0404.
Received March 31, 1998.
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References |
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