Martin Metzger

Maternal separation followed by early social deprivation affects the development of monoaminergic fiber systems in the medial prefrontal cortex of Octodon degus

The influence of early postnatal socio-emotional deprivation on the development of tyrosine hydroxylase- and 5-hydroxytryptamine-immunoreactive fiber innervation in the medial prefrontal cortex was quantitatively investigated in the precocial rodent Octodon degus. Forty-five-days-old degus from two groups were compared: (i) degus which were repeatedly separated from their mothers during the first three postnatal weeks and after weaning reared in complete isolation; and (ii) degus which were reared under normal undisturbed social conditions. The two monoaminergic fiber systems in the four subregions of the medial prefrontal cortex responded differentially to the deprivation. While the infralimbic cortex was the only subregion that displayed an increase in 5-hydroxytryptamine-positive fiber densities (129.2%) but no changes in tyrosine hydroxylase-immunoreactive fibers, the precentral medial (82.2%), anterior cingulate (74.6%) and prelimbic cortex (86.9%) showed significantly reduced tyrosine hydroxylase-positive fiber innervation, but no changes in 5-hydroxytryptamine-immunoreactive fiber densities. The number of tyrosine hydroxylase-positive somata in the ventral tegmental area and in the substantia nigra remained unchanged. In cortical areas the number of tyrosine hydroxylase-immunoreactive somata was increased (depending on the medial prefrontal cortex subregion between 241.8% and 398.7%) in deprived animals. This altered balance between the serotonergic and dopaminergic cortical innervation in the different subregions of the medial prefrontal cortex may reflect a counter-regulative anatomical and functional adaptation, which may be triggered by an altered activity of these transmitter systems during the phases of maternal separation and social isolation.

Organization of the dopaminergic innervation of forebrain areas relevant to learning: a combined immunohistochemical/retrograde tracing study in the domestic chick.

The mediorostral neostriatum/hyperstriatum ventrale (MNH) and neostriatum dorsocaudale (Ndc) of the domestic chick are crucially involved in auditory filial imprinting, whereas the lobus parolfactorius (LPO) seems to be involved in the emotional modulation of behavior. Because there is evidence that MNH and Ndc are akin to higher association areas in mammals, the present study evaluates the dopaminergic and thalamic input to these areas, as well as to the avian caudate/putamen homologue LPO, by using retrograde pathway tracing, together with dopamine (DA) and tyrosine hydroxylase (TH) immunohistochemistry. By combining DA immunohistochemistry with retrograde fluorescent tracing, we demonstrated that dopaminergic afferents to the MNH and Ndc arise mainly from the area ventralis, whereas the main dopaminergic input to the LPO arises from the substantia nigra. The main thalamic input to the MNH and LPO arises from the dorsal thalamic nuclei, n. dorsomedialis anterior and n. dorsolateralis anterior, whereas the thalamic input to the Ndc arises from the n. dorsolateralis posterior and n. subrotundus. Furthermore, there are reciprocal intratelencephalic connections between distinct parts of the neostriatum caudale and the mediorostral neostriatum. DA‐immunoreactive (ir) fibers are present at moderate densities in the MNH and Ndc and at high densities in the LPO. At the ultrastructural level, DA‐ and TH‐ir axon terminals in the MNH and Ndc form predominantly symmetric synaptic contacts with dendritic shafts, which are often situated in close vicinity to unstained terminals. These results indicate that the general organization of dopaminergic afferents to the chick telecephalon is similar to that of the mesotelencephalic dopaminergic subsystems in mammals such as the mesostriatal and mesolimbocortical DA system.

Organization of the dorsocaudal neostriatal complex: A retrograde and anterograde tracing study in the domestic chick with special emphasis on pathways relevant to imprinting

In the forebrain of domestic chicks, a network of distinct regions is crucially involved in auditory and visual filial imprinting. Among these areas, a distinct part of the dorsocaudal neostriatal complex (dNC complex), termed neostriatum dorsocaudale (Ndc), was recently discovered by its enhanced metabolic activity during the presentation of auditory and visual imprinting stimuli. Since there is evidence that the dNC complex consists of several distinct functional subareas, we investigated the neural connections of different parts of the dNC complex by retro‐ and anterograde pathway tracing. Special emphasis was put on the connections of the dNC complex with other imprinting relevant regions in the rostral telencephalon, such as the mediorostral neostriatum/hyperstriatum ventrale (MNH) and the intermediate and medial part of the hyperstriatum ventrale (IMHV). By anterograde and multiple retrograde pathway tracing, we found that the dNC complex may at least be subdivided into three major constituents. The most medial part of the dNC complex, termed neostriatum dorsale (Nd), is characterized by strong reciprocal connections with the neostriatal part of the MNH and by its auditory related inputs, including those from the output layers L1 and L3 of field L, and the shell region of the thalamic n. ovoidalis. The Ndc, which occupies the central aspects of the dNC complex, is mainly characterized by reciprocal connections with the ectostriatal belt (Ep) and the adjacent neostriatum (N). Furthermore, Nd and Ndc receive strong thalamic input from the n. dorsolateralis posterior (DLP), both project to the IMHV, and both are reciprocally connected with the archistriatum intermedium (AI). The most lateral aspect of the dNC complex, termed Ndl, is characterized by afferents from the neostriatum frontale, pars trigeminalis (NFT), and by the lack of a thalamic input. Results indicate that the dNC complex comprises distinct subregions, which are characterized by their specific afferents from parasensory areas of different sensory modalities. These different subregions may be integral components of a general pattern of sensory processing in the avian telencephalon. The strong interconnections between Nd, Ndc, and MNH as well as IMHV may constitute essential parts of auditory and visual imprinting circuits. J. Comp. Neurol. 395:380–404, 1998.

Differential projections from the lateral habenula to the rostromedial tegmental nucleus and ventral tegmental area in the rat.

The mesopontine rostromedial tegmental nucleus (RMTg) is a mostly γ‐aminobutyric acid (GABA)ergic structure believed to be a node for signaling aversive events to dopamine (DA) neurons in the ventral tegmental area (VTA). The RMTg receives glutamatergic inputs from the lateral habenula (LHb) and sends substantial GABAergic projections to the VTA, which also receives direct projections from the LHb. To further specify the topography of LHb projections to the RMTg and VTA, small focal injections of the anterograde tracer Phaseolus vulgaris leucoagglutinin were aimed at different subdivisions of the LHb. The subnuclear origin of LHb inputs to the VTA and RMTg was then confirmed by injections of the retrograde tracer cholera toxin subunit b into the VTA or RMTg. Furthermore, we compared the topographic position of retrogradely labeled neurons in the RMTg resulting from VTA injections with that of anterogradely labeled axons emerging from the LHb. As revealed by anterograde and retrograde tracing, LHb projections were organized in a strikingly topographic manner, with inputs to the RMTg mostly arising from the lateral division of the LHb (LHbL), whereas inputs to the VTA mainly emerged from the medial division of the LHb (LHbM). In the RMTg, profusely branched LHb axons were found in close register with VTA projecting neurons and were frequently apposed to the latter. Overall, our findings demonstrate that LHb inputs to the RMTg and VTA arise from different divisions of the LHb and provide direct evidence for a disynaptic pathway that links the LHbL to the VTA via the RMTg. J. Comp. Neurol. 520:1278–1300, 2012.

Localization of dopamine D1 receptors and dopaminoceptive neurons in the chick forebrain

The distributions of dopamine D1 receptors, dopaminoceptive neurons, and catecholaminergic fibers were investigated in the forebrain of the domestic chick by using D1 receptor autoradiography and immunohistochemical detection of D1 receptor protein (D1rp), the dopamine‐ and cAMP‐regulated phosphoprotein DARPP‐32, and tyrosine hydroxylase (TH). Particular attention was paid to two forebrain regions, the mediorostral neostriatum/hyperstriatum ventrale (MNH) and neostriatum dorsocaudale (Ndc), which have been shown to be crucially involved in filial imprinting. In general, there was a good, but not complete, correlation between the immunohistochemical pattern of DARPP‐32 positive perikarya and the distribution of D1 receptors. Both, DARPP‐32 positive neurons as well as D1 receptors were highly enriched in the striatal part of the basal ganglia including the lobus parolfactorius (LPO) and paleostriatum augmentatum. High to moderate densities were observed in the outer rind of the pallium. Low to moderate densities were found in the belt regions of primary sensory areas, whereas densities in the respective core regions were generally low. Labeling in the MNH and Ndc was heterogeneous. Whereas the neostriatal part of MNH displayed both, moderate DARPP‐32 immunostaining and moderate D1 receptor densities, the hyperstriatal part showed also moderate D1 receptor densities but was only weakly labeled by DARPP‐32. The rostral part of the Ndc was among the most intensely DARPP‐32 labeled areas of the pallium, its caudal part revealed only moderate DARPP‐32 immunostaining. By using D1 receptor autoradiography, a homogeneous labeling throughout the rostrocaudal extension of the Ndc was found. Double‐labeling experiments with antibodies to DARPP‐32 and TH revealed that TH positive fibers in the MNH, Ndc, and LPO were often closely related to DARPP‐32 positive perikarya. At the ultrastructural level, both immunoreaction for D1rp and DARPP‐32 in the MNH and Ndc were primarily found to be associated with postsynaptic elements. Whereas D1rp immunoreactivity was enriched at postsynaptic densities or in their vicinity, reaction product for DARPP‐32 was present throughout the perikaryal cytoplasm, dendrites, and dendritic spines. These results indicate that DARPP‐32 as well as D1 receptors in the avian forebrain reveal a distribution that is substantially similar to that of mammals. J. Comp. Neurol. 388:146–168, 1997.

Lateral habenula and the rostromedial tegmental nucleus innervate neurochemically distinct subdivisions of the dorsal raphe nucleus in the rat

The lateral habenula (LHb) is an epithalamic structure differentiated in a medial (LHbM) and a lateral division (LHbL). Together with the rostromedial tegmental nucleus (RMTg), the LHb has been implicated in the processing of aversive stimuli and inhibitory control of monoamine nuclei. The inhibitory LHb influence on midbrain dopamine neurons has been shown to be mainly mediated by the RMTg, a mostly GABAergic nucleus that receives a dominant input from the LHbL. Interestingly, the RMTg also projects to the dorsal raphe nucleus (DR), which also receives direct LHb projections. To compare the organization and transmitter phenotype of LHb projections to the DR, direct and indirect via the RMTg, we first placed injections of the anterograde tracer Phaseolus vulgaris leucoagglutinin into the LHb or the RMTg. We then confirmed our findings by retrograde tracing and investigated a possible GABAergic phenotype of DR‐projecting RMTg neurons by combining retrograde tracing with in situ hybridization for GAD67. We found only moderate direct LHb projections to the DR, which mainly emerged from the LHbM and were predominantly directed to the serotonin‐rich caudal DR. In contrast, RMTg projections to the DR were more robust, emerged from RMTg neurons enriched in GAD67 mRNA, and were focally directed to a distinctive DR subdivision immunohistochemically characterized as poor in serotonin and enriched in presumptive glutamatergic neurons. Thus, besides its well‐acknowledged role as a GABAergic control center for the ventral tegmental area (VTA)–nigra complex, our findings indicate that the RMTg is also a major GABAergic relay between the LHb and the DR. J. Comp. Neurol. 522:1454–1484, 2014.

Prefrontal afferents to the dorsal raphe nucleus in the rat

The prefrontal cortex (PFC) receives strong inputs from monoaminergic cell groups in the brainstem and also sends projections to these nuclei. Recent evidence suggests that the PFC exerts a powerful top-down control over the dorsal raphe nucleus (DR) and that it may be involved in the actions of pharmaceutical drugs and drugs of abuse. In the light of these findings, the precise origin of prefrontal inputs to DR was presently investigated by using the cholera toxin subunit b (CTb) as retrograde tracer. All the injections placed in DR produced retrograde labeling in the medial, orbital, and lateral divisions of the PFC as well as in the medial part of the frontal polar cortex. The labeling was primarily located in layer V. Remarkably, labeling in the medial PFC was denser in its ventral part (infralimbic and ventral prelimbic cortices) than in its dorsal part (dorsal prelimbic, anterior cingulate and medial precentral cortices). After injections in the rostral or caudal DR, the largest number of labeled neurons was observed in the medial PFC, whereas after injections in the mid-rostrocaudal DR, the labeled neurons were more homogeneously distributed in the three main PFC divisions. A cluster of labeled neurons also was observed around the apex of the rostral pole of the accumbens, especially after rostral and mid-rostrocaudal DR injections. Overall, these results confirm the existence of robust prefrontal projections to DR, mainly derived from the ventral part of the medial PFC, and underscore a substantial contribution of the frontal polar cortex.

Possible crosstalk between leptin and prolactin during pregnancy

Rodents exhibit leptin resistance and high levels of prolactin/placental lactogens during pregnancy. A crosstalk between prolactin and leptin signaling has been proposed as a possible mechanism to explain the changes in energy balance during gestation. However, it remains unclear if specific neuronal populations co-express leptin and prolactin receptors. Therefore, our present study was undertaken to identify in the mouse brain prolactin-responsive cells that possibly express the leptin receptor (LepR). In addition, we assessed the leptin response in different brain nuclei of pregnant and nulliparous mice. We used a LepR-reporter mouse to visualize LepR-expressing cells with the tdTomato fluorescent protein. Prolactin-responsive cells were visualized with the immunohistochemical detection of the phosphorylated form of the signal transducer and activator of transcription-5 (pSTAT5-ir). Notably, many neurons that co-expressed tdTomato and pSTAT5-ir were observed in the medial preoptic area (MPA, 27-48% of tdTomato cells), the retrochiasmatic area (34-51%) and the nucleus of the solitary tract (NTS, 16-24%) of prolactin-treated nulliparous mice, pregnant mice and prolactin-treated leptin-deficient (ob/ob) mice. The arcuate nucleus of the hypothalamus (8-22%), the medial tuberal nucleus (11-15%) and the ventral premammillary nucleus (4-10%) showed smaller percentages of double-labeled cells among the groups. Other brain nuclei did not show significant percentages of neurons that co-expressed tdTomato and pSTAT5-ir. Late pregnant mice exhibited a reduced leptin response in the MPA and NTS when compared with nulliparous mice; however, a normal leptin response was observed in other brain nuclei. In conclusion, our findings shed light on how the brain integrates the information conveyed by leptin and prolactin. Our results corroborate the hypothesis that high levels of prolactin or placental lactogens during pregnancy may directly interfere with LepR signaling, possibly predisposing to leptin resistance.

Thalidomide treatment down-regulates SDF-1α and CXCR4 expression in multiple myeloma patients

The chemokine stromal-derived factor-1alpha (SDF-1alpha) and its receptor CXCR4 are critically involved in directional migration and homing of plasma cells in multiple myeloma. Here, we show that the expression of SDF-1alpha and CXCR4 was significantly down-regulated in patients treated with thalidomide (n=10) as compared to newly diagnosed MM patients (n=31) and MM patients treated with other drugs (n=38). SDF-1 alpha and CXCR4 expression was also significantly decreased in a RPMI 8226 cell line treated with 10 and 20micromol/L of thalidomide. Our findings indicate that thalidomide therapy induces down-regulation of CXCR4 and its ligand SDF-1alpha in multiple myeloma.

Serotonergic innervation of the telencephalon in the domestic chick

The serotonergic system in the telencephalon of the domestic chick was investigated using an antibody specific to serotonin (5-HT). Most parts of the forebrain, such as the different subdivisions of the visual Wulst and the neostriatum, displayed a rather uniform, moderate to dense innervation of serotonergic (5-HT+) fibers. However, some highly distinct area-specific differences could be observed. Primary sensory areas such as the ectostriatum, layer L2 of field L, and the rostral part of the nucleus basalis displayed very few 5-HT+ fibers. In contrast, the dorsal part of the archistriatum intermedium, the nucleus taeniae, a medial part of the lobus parolfactorius and the dorsomedial part of the hippocampus displayed an extremely dense serotonergic innervation. In general, three different types of 5-HT+ axons could be distinguished. The most common was a fine, highly varicose type, whereas beaded axons, exhibiting larger varicosities, and a thick non-varicose type, exhibiting occasional swellings, were much sparser. In summary, these findings indicate that the serotonergic innervation of the avian telencephalon is extensive but site-specific, and is organized in a highly similar way to that in mammals. The high accumulation of 5-HT+ fibers in the dorsal part of the archistriatum intermedium points to a prominent role for 5-HT in fear behavior.