Prevention
by morphine of N-Methyl-D-Aspartic Acid induced
penile erection and yawning: involvement of
Nitric Oxide
MR Melis, S Succu, A Argiolas
Bernard B. Brodie Department
of Neuroscience, University of Cagliari,
Italy
Introduction : Penile erection and yawning
are two different behavioral patterns that often
occur concoinitantly in different experimental
and physiological conditions. Among substances,
that induce both penile erection and yawning,
the best known are dopamine receptor agonists
(i.e., apomorphine), oxytocin,
adrenocorticotropin, and related peptides, 5-HT
receptor agonists that act mainly ou the 5-HT1c
receptor subtype and N-methyl-D-aspartic acid
(NMDA). lnterestinlgly, NMDA-induced penile
erection and yawning are apparently mediated by
the stimulation of the excitatory aminoacid
receptors of the NMDA subtype because they are
prevemed by dizolcipine (MK-801), a
noncompetitive antagonist of these receptors,
and are not induced by
(-)-a-(amino)-3-hydroxy-5-methylisoxazole-4propionic
acid (AMPA) or by
trans-l-amino-1,3-cyclopentanedicarboxylic acid
(ACPD), agonists of the AMPA and metabotropic
receptor subtype, respectively. NMDA, like
dopamine agonists and oxytocin, induces penile
erection and yawning by acting in the
paraventricular nucleus of the hypothalamus
(PVN), apparently by activaling central
oxytocinergic transmission, these being
responses prevented by the central
administration of oxytocin receptor
antagonists.
Experimental evidence also suggest that
NMDA, like dopamine agonists and oxytocin,
induces penile erection and yawning by
activating nitric oxide (NO) synthase in the
PVN, which in turn leads to an increase of
central oxytocinergic transmission. This
supports the involvement of the recently
discovered neurotransniitter/neuromodulator NO
in the expression of penile erection and yawning
induced by NMDA.
Accordingly, NO donors injected into the PVN
induce penile erection and yawning that are
indistinguishable from those induced by NMDA and
that are prevented by the potent oxytocin
receptor antagonist
d(CH2)5Tyr(Me)-Orn8-vasotocin -given into the
lateral ventricles (ICV). The involvement of NO
in the induction of penile crection and yawning
by NMDA has been recently substantiated by
measuring the production of NO in vivo in the
PVN of male rats by microdialysis. Indeed,
NMDA-induced penile erection and yawning were
found to be associated to an increase of the
concentration of NO2 and NO3, the reaction
products of newly synthetized NO with O2 that
are an indirect but reliable indicator of NO
production by biological tissues in vivo, in the
dialysate collected from a vertical
inicrodialysis probe implanted in the PVN.
Furthermore, this NMDA-induced NO2 and NO3
increase was prevented not only by MK-801 as
expected, but also by NO synthase inhibitors
given ICV or into the PVN, which prevented also
NMDA-induced penile crection and yawning. This
report shows that these NMDA-induced behavioral
responses are also prevented by the opiate
morphine injected in the PVN and that this
effect of the opiate on penile erection and
yawning induced by NMDA is related to a
concomitant prevention of the NO production
induced by NMDA in tnePVN of male rats as
determined by in vivo microdialysis.
[...]
Discussion : The present results show
that morphine injected in the PVN antagonizes
dose dependently penile erection and yawning
induced by NMDA injected ni the paraventricular
nucleus of the hypothalamus. Most important,
this inhibitory effect of morphine on
NMDA-induced penile crection and yawning was
related to a concomitant prevention of the
NMDA-induced increase of the NO2 and NO3
concentration in the paraventricular dialysate.
This finding not only adds further support to
the hypothesis that NMDA induces penile erection
and yawning by stimulating NO synthase in the
PVN, as found in several other brain areas, but
also suggests that morphine prevents NMDA
responses by preventing NMDA-induced NO synthase
activation. Indeed. the concentration of NO2 and
NO3- in the extracellular fluids represents a
reliable indicator of the production of NO in
biological ttissues, that is of the activity of
NO synthase, which converts L-arginine to
citrulline and NO at least when blood is absent.
In this condition. the newly formed NO would be
oxidized in few seconds by O2, mainly to NO2
and, to a lesser extent, to NO31. The effect of
morphine on NMDA-induced increase of NO
production, penile erection. and yawning seems
to be mediated by the stimulation of opioid
receptors, because these effects are prevented
by the prior blockade of opioid receptors by
naloxone. The reccptor involved in mediating
morphine effect seems to be of the µ
subtype, because U-69,593, an opioid agonist 500
times more selective than morphine on the opioid
receptors of the k subtype is ineffective on
NMDA responses.
The prevention by morphine of NMDA-induced
paraventricular NO production, penile erection,
and yawning resembles the ability of the opiate
to prevent these responses when induced by
apomorphine and by oxytocin injected in the PVN.
Because NMDA-induced penile erection and yawning
are also prevented hN the ICV administration of
potent oxytocin receptor anta-onists 121 j~
which also prevent these behavioral responses
when induced by apomorphine and oxytocin, the
findings are in fine with the hypothesis that
opioid receptors in the PVN control in an
inhibitory fashion those oxytocinergic neurons
projecting to extrahypothalainic brain areas
whose activation by NMDA, dopamine or oxytocin
leads to the expression of penile erection and
yawning by activating NO synthase in the PVN.
Accordingly, inhibitory µ opioid receptors
are present in the cell bodies of
paraventricular oxytocinergic neurons projecting
to the neurohypophysis.
However, it must be noted that the
prevention by morphine of the behavioral
responses induced by NMDA was observed at doses
of the opiate that did not prevent a completely
NMDA-induced increase of the concentration of
NO2 and NO3 in the paraventricular dialysate. A
possible explanation for this discrepancy is
that NMDA also activates NO synthase in neurons
that are not involved in the expression of
penile erection and yawing. In agreement with
this possibility, NO synthase has been
identified not only in oxytocinergic but also in
vasopressinergic cell bodies, in the PVN. If one
assumes that NMDA activation of NO synthase is
secondary to an increased Ca 2+ influx through
the NMDA receptor coupled Ca2+ channels, as
suggested in the cerebellum and the hippocampus,
a possible rnechanism by means of which morphine
prevents the NMDA induced activation of NO
synthase in the PVN, is that the opiate
decreases Ca 2+ influx, which in turn causes a
decrease in the activity of NO synthase, a
calcium-calmodulin-dependent enzyme [ in the
cell bodies of oxytocinergic neurons mediatig
penile erection and yawning.
The decreased NO formation would, in turn,
reduce the oxytocinergic transmission by a yet
unknown mechanism, apparently unrelated to the
activation of guanylate cyclase, thereby
decreasing the behavioral responses.
Accordingly, the inhibition of Ca2+ influx by
stimulation of opioid receptors coupled to G
proteins or directly to Ca2+ channels has been
already reported in several tissues and cells,
and the prevention by morphine of NMDA-induced
penile erection and yawning resembles the
prevention of these responses induced by
apomorphine and oxytocin by the blockade of
N-type voltage-dependent calcium channels by
nanogram amounts of omega-conotoxin injected
into the PVN.
However, the present results do not rule out
other mechanisms by means of which morphine
mlght interfere with NO synthase activation. For
instance, morphine might alter NO synthase
activity in oxytocinergic neurons by modulating
the release of other
neurotransmitters/neuromodulators in the PVN or
the activity of adenylate cyclase, one of the
best-knwon target of opioid peptides.
In contrast, a direct inhibition of NO
synthase by morphine is unlikely, because (1)
morphine alone does not cause any decrease in
basal NO2 and NO3- levels (this study), and (2)
acute or chionic morphine does not alter NO
synthase activity ni brain homogenates or its
expression, as measured by the amount of
NO-synthase mRNA in several mouse brain
areas.
In contrast with the results of this study
and of other studies stimulatory effects of
morphine ou NO synthase activity have been
reported. This morphine effect bas been found in
homogenates of mouse cerebellum but not of other
brain areas in cultured human endothelial cells,
and invertebrate microglia. The reason for this
discrepancy is unknown. One possibility is that
the receptors, activated by morphine in these
tissues are linked to transduction systems
different from those present in the PVN.
Accordingly, in endothelial cells and
invertebrate microglia morphine was known to
induce a short-lasting NO release by acting on a
so-called µ3 receptor, which is sensitive
to morphine but not to opioid peptides and which
has not been identified yet in brain.
In this regard it is noteworthy that
morphine inhibits drug induced penile erection
and yawning through opioid peptide-sensitive
µ receptors, because stable enkephalin and
P-endorphin analogs are as effective as morphine
in preventing, these beliavioral response.
Interactions between opioids and excitalory
aniino acids and opioids and NO have been shown
to occur in other central functions. For
instance, opioid antinociception, tolerance and
dependence can be antagonized by excitatory
amino acid receptor antagonists, suggesting that
opioids and glutamic acid can act
sinergistically at least in the control of the
above functions. Likewise, NO seems to be
involved in the expression of opioïd
antinociception tolerance and dependence
although with some discrepancy. In these studies
either facilitatory or inhibitory effects of NO
or morphine effects were find. In conclusion,
the present results show that morphine interacts
with glutamic acid and NO in opposite manner in
the control of penile erection and yawing at the
level. In particular, these findings raise the
possibility that µ opioid receptors might
decrease NO synthase activity through machanisms
related to the closure/opening of ion channels
thate infere withe Ca2+ influx. Although the
mechanism of this inetraction has still to be
elucidated, our results confirm and extend
precious studies that µ opiod receptors
exert potent inhibitory effects on penile
erection and yawning, and that these effects are
in part mediated by the inhibition of NO
synthase in the PVN.
