MePD Neurons Research Articles

Sexual dimorphism, estrous cycle and laterality determine the intrinsic and synaptic properties of medial amygdala neurons in rat.

The posterodorsal medial amygdala (MePD) is a sex steroid-sensitive area that modulates different social behavior by relaying chemosensorial information to hypothalamic nuclei. However, little is known about MePD cell type diversity and functional connectivity. Here, we have characterized neurons and synaptic inputs in the right and left MePD of adult male and cycling female (in diestrus, proestrus or estrus) rats. Based on their electrophysiological properties and morphology, we found two coexisting subpopulations of spiny neurons that are sexually dimorphic. They were classified as Class I (predominantly bitufted-shaped neurons showing irregular spikes with frequency adaptation) or Class II (predominantly stellate-shaped neurons showing full spike frequency adaptation). Furthermore, excitatory and inhibitory inputs onto MePD cells were modulated by sex, estrous cycle and hemispheric lateralization. In the left MePD, there was an overall increase in the excitatory input to neurons of males compared to cycling females. However, in proestrus, the MePD neurons received mainly inhibitory inputs. Our findings indicate the existence of hemispheric lateralization, estrous cycle and sexual dimorphism influences at cellular and synaptic levels in the adult rat MePD.

Methamphetamine and Ovarian Steroid Responsive Cells in the Posteriodorsal Medial Amygdala are Required for Methamphetamine-enhanced Proceptive Behaviors

Methamphetamine (Meth) is a psychomotor stimulant strongly associated with increases in sexual drive and impulse in both men and women. These changes in sexual motivation have a greater impact on women due to their likelihood of facing the greater burden of unplanned pregnancies, as well as increased risk for psychiatric co-morbidities such as depression. We have previously established a rodent model of Meth-induced increases in sexual motivation. Using this model, we have identified the posteriodorsal medial amygdala (MePD) via excitotoxic lesion studies as a necessary nucleus in Meth-facilitated female sexual motivation. While lesion studies give us insight into key nuclei that may be targets of Meth action, such an approach does not give insight into the identity of the specific MePD neurons or neural circuitry involved in Meth-induced increases in proceptive behaviors. Using the DAUN02 inactivation method, a recently established technique for removing behaviorally relevant cell populations, we present evidence that the ovarian steroid/Meth responsive cells in the MePD are necessary for Meth-induced facilitation of proceptive behaviors. These findings form the basis for future work that will allow for the classification of neuronal subtypes involved in the MePD’s modulation of proceptive behavior as well as a stronger understanding of the neurocircuitry of female sexual motivation.

Castration alters the number and structure of dendritic spines in the male posterodorsal medial amygdala.

The posterodorsal medial amygdala (MePD) is responsive to androgens and participates in the integration of olfactory/vomeronasal stimuli for the display of sexual behavior in rats. Adult gonadectomy (GDX) affects the MePD structural integrity at the same time that impairs male mating behavior. At the cellular level, dendritic spines modulate excitatory synaptic transmission, strength, and plasticity. Here, we describe the effect of GDX on the number and shape of dendritic spines in the right and left MePD using confocal microscopy and 3D image reconstruction. Age-matched adult rats were intact (n=6), submitted to a sham procedure (n=4) or castrated and studied 90days after GDX (n=5). The MePD neurons have a density of 1.1 spines/dendritic μm composed of thin, mushroom-like, stubby/wide, and few ramified or atypical spines. Irrespective of brain hemisphere, GDX decreased the dendritic spine density in the MePD, but induced different effects on each spine type. That is, compared to control groups, GDX reduced (i) the number (up to 50%) of thin, mushroom-like, and ramified spines, and (ii) the size and the neck length of thin spines as well as the head diameter of ramified spines. Besides, GDX increased the number of stubby/wide and atypical spines (up to 140% and 400%, respectively). These data show that GDX promotes a cellular and synaptic reorganization in a spine-specific manner in the MePD. By altering the number and shape of these connectional elements, GDX can affect the neural transmission and hinder the function of integrated brain circuitries in the male brain.

Inhibitory and multisynaptic spines, and hemispherical synaptic specialization in the posterodorsal medial amygdala of male and female rats

The density of dendritic spines is sexually dimorphic and variable throughout the female estrous cycle in the rat posterodorsal medial amygdala (MePD), a relevant area for the modulation of reproductive behavior in rats. The local synaptic activity differs between hemispheres in prepubertal animals. Here we used serial section transmission electron microscopy to produce 3D reconstructions of dendritic shafts and spines to characterize synaptic contacts on MePD neurons of both hemispheres in adult males and in females along the estrous cycle. Pleomorphic spines and nonsynaptic filopodia occur in the MePD. On average, 8.6% of dendritic spines received inputs from symmetric gamma-aminobutyric acid (GABA)-immunoreactive terminals, whereas 3.6% received two synaptic contacts on the spine head, neck, or base. Presynaptic terminals in female right MePD had a higher density of synaptic vesicles and docked vesicles than the left MePD, suggesting a higher rate of synaptic vesicle release in the right MePD of female rats. In contrast, males did not show laterality in any of those parameters. The proportion of putative inhibitory synapses on dendritic shafts in the right MePD of females in proestrus was higher than in the left MePD, and higher than in the right MePD in males, or in females in diestrus or estrus. This work shows synaptic laterality depending on sex and estrous cycle phase in mature MePD neurons. Most likely, sexual hormone effects are lateralized in this brain region, leading to higher synaptic activity in the right than in the left hemisphere of females, mediating timely neuroendocrine and social/reproductive behavior.

Descriptive findings on the morphology of dendritic spines in the rat medial amygdala

The rat posterodorsal medial amygdala (MePD) is a brain area in which gonadal hormones induce notable plastic effects in the density of dendritic spines. Dendritic spines are post-synaptic specializations whose shape and spacing change neuronal excitability. Our aim was to obtain new data on the dendritic spines morphology and density from MePD neurons using the carbocyanine dye DiI under confocal microscopy. In adult male rats, the dendritic spine density of the medial branches of the left MePD (mean ± SD) was 1.15 ± 0.67 spines/dendritic μm. From the total sampled, approximately 53% of the spines were classified as thin, 22.5% as “mushroom-like”, and 21.5% as stubby/wide. Other spine shapes (3%) included those ramified, with a filopodium-like or a gemule appearance, and others with a protruding spinule. Additional experiment joining DiI and synaptophysin (a pre-synaptic protein) labeling suggested synaptic sites on dendritic shafts and spines. Dendritic spines showed synaptophysin puncta close to their head and neck, although some spines had no evident labeled puncta on them or, conversely, multiple puncta appeared upon one spine. These results advance previous light microscopy results by revealing features and complexities of the dendritic spines at the same time that give new insight on the possible synaptic organization of the adult rat MePD.

The density of Golgi-impregnated dendritic spines from adult rat posterodorsal medial amygdala neurons displays no evidence of hemispheric or dorsal/ventral differences

The posterodorsal medial amygdaloid nucleus (MePD) is a sexually dimorphic area in the rat brain and dendritic spines are specialized postsynaptic sites involved with local neural plasticity. Previous electrophysiological data showed that prepubertal males have more excitatory synapses than females in the left MePD. Besides, dorsal and ventral MePD neurons have a heterogeneous expression of estrogen receptors α or β in mating-responsive neurons in females. Based on these findings, the “single-section” Golgi method was employed in adult rats ( n = 6 in each group) to reveal: (1) the effect of hemispheric laterality in the density of dendritic spines in the MePD of males and diestrus females, and (2) the density of dendritic spines in the MePD dorsal and ventral subregions in proestrus females (mean values from n = 48 neurons for each experimental variable). There were no statistically significant differences for sex, laterality or the interaction of these factors in the dendritic spine density between males and diestrus females ( p > 0.2), nor for the dorsal and the ventral MePD dendritic spine density in proestrus females ( p > 0.1). These findings complement current knowledge about the rat MePD and suggest that the number of proximal dendritic spines is not lateralized at adulthood. Furthermore, the differential expression of estradiol receptors in the dorsal and ventral MePD did not lead to distinct spine number in these subregions when circulating ovarian steroids peak in proestrus.

Efferent Projections of the Anterior and Posterodorsal Regions of the Medial Nucleus of the Amygdala in the Mouse

The efferent projections of the anterior and posterodorsal part of the medial nucleus (MePD) in the mouse were studied by means of anterograde axonal tracing using biotinylated dextran amine. The MePD axons ran mainly via the stria terminalis and to a lesser extent via the ventral amygdalofugal pathway. The projections to the forebrain were broadly distributed and varied from very strong to scant. The most significant connections were destined to the bed nucleus of the stria terminalis in which all parts of the medial division were innervated by MePD neurons. Moderate projections reached the limbic striatum (nucleus accumbens), olfactory tubercle and the lateral septal nucleus. The substantia innominata was also innervated by the MePD, and especially the projection to its ventral portion was substantial. The profuse innervation of the medial preoptic nucleus and medial preoptic area indicated significant involvement of the MePD in sexual behavior. Many hypothalamic nuclei were innervated but to a different extent. The very strong innervation of the ventral premammillary nucleus further indicated the involvement of the MePD in the neuronal circuitry for sexual behavior. Substantial projections also reached the anterior hypothalamus and tuber cinereum, while the connections to the lateral hypothalamus were widespread but showed moderate density. MePD strongly innervated the ventrolateral part of the ventromedial hypothalamic nucleus and moderately its remaining parts. The neurosecretory hypothalamic nuclei and the arcuate nucleus contained only a few MePD terminals. The thalamic innervation was very scant and reached the lateral habenular nucleus and the nuclei of the midline. The mesencephalic connections were moderate to sparse and projected to the mesolimbic dopaminergic groups in the ventral tegmental area, the pars lateralis and the dorsal tier of the substantia nigra pars compacta, the periaqueductal gray and the dorsal raphe nucleus. The present results principally resembled data known in other rodent species; however, the efferents of the MePD often differed in extent and/or topical distribution.

Dendritic spine density of posterodorsal medial amygdala neurons can be affected by gonadectomy and sex steroid manipulations in adult rats: A Golgi study

The posterodorsal medial amygdala (MePD) is a sex steroid-responsive area in the rat brain. The dendritic spine density of Golgi-impregnated MePD neurons were studied in: (1) adult gonadectomized (GDX) males after a short or a longer postcastration period (8 and 90 days, respectively), compared to age-matched sham operated and to intact controls; (2) adult GDX females, which received oil, estradiol benzoate (EB) alone or EB and progesterone as substitutive therapy; and, (3) EB-treated GDX females that concomitantly received saline or LY235959, a competitive antagonist of NMDA receptors, to test a possible glutamatergic mediation on the estrogen-mediated increase in spine density in this brain area. Intact males showed a higher spine density than males studied 8 days after sham operation or those in both short- and long-term GDX groups ( p < 0.02), but not when compared to males at 90 days after sham operation ( p = 0.12). In females, dendritic spine density increased following EB injections when compared to the oil group ( p = 0.05), with an effect that was potentiated by progesterone ( p < 0.01). LY235959 was not able to block the stimulating effect of EB on dendritic spines of GDX females ( p > 0.2). These data provide novel evidence that MePD dendritic spines are affected by sex steroid manipulations in adult rats, GDX males had a specific spine density decrease after a long postcastration period, and estrogen (apparently independently of a NMDA receptor interaction) and progesterone have stimulatory effects on the number of dendritic spines in GDX females.

Sexual dimorphism in neuronal number of the posterodorsal medial amygdala is independent of circulating androgens and regional volume in adult rats

The posterodorsal medial amygdala (MePD) in rodents integrates olfactory and pheromonal information, which, coupled with the appropriate hormonal signals, may facilitate or repress reproductive behavior in adulthood. MePD volume and neuronal soma size are greater in male rats than in females, and these sexual dimorphisms are maintained by adult circulating hormone levels. Castration of adult males causes these measures to shrink to the size seen in females 4 weeks later, whereas testosterone treatment of adult females for 4 weeks enlarges these measures to the size of males. We used stereological methods to count the number of cells in the MePD and found that, in addition to the sex difference in regional volume and soma size, males also have more MePD neurons than do females, yet these numbers are unaffected by the presence or absence of androgen in adults of either sex. Males also have more glial cells than do females, but, in contrast to the effects on neuronal number, the number of glial cells is affected by androgen in the right MePD of both sexes and, therefore, may contribute to regional volume changes in adulthood in that hemisphere. Thus, regional volume, neuronal size, and glial numbers vary in the MePD of adult rats in response to circulating androgens, but neuronal number does not. These results suggest that the sex difference in neuronal number in the rat MePD may be "organized" by androgens prior to adulthood, whereas regional volume, neuronal size, and glial numbers can be altered by androgens in adulthood.

Dendritic branching features of posterodorsal medial amygdala neurons of adult male and female rats: Further data based on the Golgi method

The posterodorsal portion of the medial amygdalar nucleus (MePD) contains receptors for gonadal hormones and modulates the function of a social behavior network in rodents. The aims of this study were: to provide further data about the morphology of Golgi-impregnated dendrites of neurons from the MePD of adult rats; and, to compare the results obtained for dendritic branching and predominant dendritic spatial distribution in the MePD of males and diestrus females. MePD neurons were classified as bitufted or stellate, their spiny dendrites showed variable lengths, divided sparingly and decreased the number of branches with the distance from the soma. Dendritic arborization levels, number of branches in each level, distribution of the dendrites around the cell body and away from it, and the preferred spatial distribution of dendritic branches were studied according to different techniques and compared between sexes. Statistically significant differences were found in the predominant dendritic spatial distribution in the MePD, males with branches more oriented medially and dorsolaterally and females with more dorsally and ventromedially ones (p< or =0.05 in all cases). This result adds another clue to understand how information is processed and integrated in the MePD and within functionally dynamic sex steroids-responsive circuits relevant for reproduction in both sexes.

Influence of sex and estrous cycle, but not laterality, on the neuronal somatic volume of the posterodorsal medial amygdala of rats

The aim of the present study was to measure the cell body volume of neurons from the posterodorsal subnucleus of the medial amygdala (MePD) of adult male ( n = 5) and diestrus, proestrus and estrus female ( n = 4–5 in each group) rats to reveal a possible sexual dimorphism, estrous cycle variations and laterality in this morphological parameter. The brains of adult Wistar rats were sectioned (1 μm), stained with 1% toluidine blue and the stereological estimation of neuronal soma volume of both sides of MePD was realized using the Cavalieri method and the technique of point counting. Data were compared by a two-way ANOVA for repeated measures and the least significance difference post hoc test. In the MePD, mean neuronal somatic volume showed a statistical difference among groups ( p = 0.005), but neither an effect of laterality ( p = 0.33) nor interactions between groups and laterality ( p = 0.78) were found. Post hoc test showed that males (mean ± S.E.M., 2075.67 ± 135.79 μm 3) have larger mean neuronal somatic volume compared to females in proestrus (1503.30 ± 44.46 μm 3) and in estrus (1616.69 ± 71.49 μm 3, p < 0.05 in both cases), but not in diestrus (1940.78 ± 129.68 μm 3, p > 0.05). Moreover, diestrus females displayed larger mean neuronal somatic volume than proestrus female rats ( p < 0.05). It is suggested that neuronal somatic volume is another sexually dimorphic finding in the MePD, for which it is relevant to set apart the different phases of the estrous cycle to reveal the presence of gonadal hormones effects in the rat MePD neurons.

Dendritic pruning of the medial amygdala during pubertal development of the male Syrian hamster

The medial amygdala (Me), a brain region essential for mating behavior, changes in size during puberty. In pre-, mid-, and late pubertal (21, 35, and 49 days of age) male Syrian hamsters, we examined neuronal structure in Me and protein levels of spinophilin and synaptophysin in the amygdaloid complex for evidence of synaptic plasticity coincident with behavioral and physiological development. Body weight, testes weight, and testosterone levels increased during puberty. Mounting behavior, including ectopic, nonintromittive, and intromittive mounts, also increased. Neuronal structure in the posterodorsal medial amygdala (MePD) was assessed in Golgi-impregnated neurons. Pruning occurred during puberty in the number of dendrites emanating from the cell body and in terminal dendritic spine densities. Approximately half of all MePD neurons analyzed had an axon emanating from a dendrite rather than the cell body. However, prepubertal males were more likely to have the axon emanating from a higher order dendritic segment (secondary or tertiary) than were mid- and late pubertal males. Finally, protein levels in the amygdaloid complex varied with pubertal age. Spinophilin decreased, while synaptophysin and GAPDH protein levels increased. These results suggest that puberty is a period of dramatic synaptic plasticity in Me. Specifically, pruning of dendrites and spines, in combination with axonal changes, is likely to modify the afferent influences and electrophysiological properties of Me neurons. Because the Me is an integral component of a social behavior neural network, these changes may be related not only to sexual behavior, but also to other behaviors that mature during puberty, including aggressive, risk-taking, fear-related, and parental behaviors.

Further studies on the rat posterodorsal medial amygdala: Dendritic spine density and effect of 8-OH-DPAT microinjection on male sexual behavior

The rat posterodorsal medial amygdala (MePD) is a component of the neural network that modulates male sexual behavior. Dendritic spines were counted in Golgi-impregnated bitufted and stellate neurons and from cells located in the medial and lateral MePD subregions. It was also studied the effect of 8-OH-DPAT, a 5-HT 1A receptor agonist, microinjected into the MePD on male sexual behavior. There were no significant differences in the dendritic spine density obtained from multipolar bitufted and stellate neurons ( n = 48 cells in each group; p > 0.05) or in the data from the medial or the lateral MePD ( n = 48 neurons per region; p > 0.05). Rats were stereotaxically microinjected into the MePD with saline (0.2 μl, n = 6) or 8-OH-DPAT (0.1 and 1.0 μg/0.2 μl, n = 6 and 5, respectively). Behavioral recordings prior to surgery and “non-target” microinjections served as additional control data. 8-OH-DPAT 1.0 μg decreased the latencies to intromission and ejaculation, the postejaculatory refractory period and the mount frequency when compared to control pre-surgery data ( p < 0.05). When compared among groups, 8-OH-DPAT 1.0 μg promoted the highest percentage reduction in the postejaculatory refractory period. Saline and injections in the vicinity of MePD did not promote relevant effects on ejaculation ( p > 0.05). Results indicate that a similar dendritic spine density can be found in morphologically different populations of MePD neurons and, 8-OH-DPAT can facilitate male sexual behavior by acting on postsynaptic 5-HT 1A receptors in this brain area.