Brainstem Afferents To The Hippocampal Formation: Comparative Inmunohistochemical Study In The Macaca Fascicularis Monkey

The neuroanatomical connections in the nonhuman primate of the brainstem structures to the Hippocampal Formation (HF, which includes the dentate gyrus -DG-, CA3, CA2, CA1, subiculum, pre-parasubiculum and the entorhinal cortex -EC-) are still unclear. Previous tracer studies in nonhuman primates show retrogradely labeled neurons in the brainstem including the Ventral Tegmental Area (VTA), Locus Coeruleus (LC) and Raphe Nuclei (RN), after deposits in the hippocampus (Amaral and Cowan, 1980), as well as in the EC (Insausti et al., 1987). In order to characterize the neurotransmitters associated to those projections (presumably dopaminergic -DA, VTA-, noradrenergic -NA, LC-, and serotoninergic -5-HT, RN-, respectively), and the topographic and laminar differences, we studied comparatively the innervation in the HF using immunohistochemical techniques. Inmunohistochemistry for DA (Tirosine Hidroxilase, TH), NA (Dopamine Beta Hidroxilase, DBH), and 5-HT showed: a) The DG molecular layer had TH-immunoreactive fibers, while the polymorphic layer contained positive 5-HT fiber labeling, b) CA3 pyramidal layer showed denser 5-HT labeling than TH, c) CA1 had scattered TH and 5-HT fibers, d) The superficial layer of the rostral EC (I and II) had TH- and 5-HT-labelled processes, e) TH and DBH positive cells were primarily found in the lateral subdivisions of the EC (ELR/ELc). The preferential location of these positive fibers in ELR/ELc, is significant, as this portion of the EC receives abundant unimodal and polymodal sensory input and innervates the body and tail of the hippocampus, and therefore it might be a crucial link in the consolidation of memory through the monoaminergic modulation of the HF

Hippocampal Formation Projection To Ventral Tegmental Area: An Anatomical Study In The Non-Human Primate

The Hippocampal Formation (HF) has a critical role in episodic memory. One of the major components in episodic memory is the encoding of novel stimuli, which is associated to dopaminergic system. Lisman and Grace (2005) proposed that novelty signals in the hippocampus modulate the activity of dopaminergic neurons in the ventral tegmental area (VTA) and that, via a feedback loop, the increase of dopamine in hippocampal neurons promotes the encoding for the novel event. Retrograde tracer studies have demonstrated that the VTA projects directly to the HF in primates (Amaral and Cowan, 1980; Insausti et al., 1987) as well as in rodents. However, whether these projections are reciprocal or not is unknown. Despite this lack of evidence of a direct projection, functional studies indicate that the dopaminergic neurons of the VTA are strongly influenced by the hippocampus indirectly through either lateral septum (Luo et al. 2011) or nucleus accumbens-ventral pallidum pathways (Lisman and Grace 2005). In order to determine the existence of direct inputs from the HF to the VTA and which are the specific fields within the HF responsible of the projection, the retrograde tracers were placed in the mesencephalic ventral and dorsal tegmentum of the Macaca fascicularis monkey, including the VTA. The retrograde cell labeling was analyzed with an epifluorescence microscope coupled to a computerized charting system. Our preliminary results showed scarce labeled cells in the HF, specifically in dorsal subiculum, and deep layers of the caudomedial portion of the entorhinal cortex (subfield EO and medialmost EI). These results clarify the functional HF-VTA loop playing a role in learning and memory, and different neuropsychiatric diseases (schizophrenia, Alzheimer´s and Parkinson's disease

Brainstem afferents to the hippocampal formation: Comparative immunohistochemical study in the Macaca fascicularis monkey

The synaptic plasticity of the Hippocampal Formation (HF, which includes the dentate gyrus -DG-, CA3, CA2, CA1, subiculum, pre-parasubiculum and the entorhinal cortex -EC) is strongly influenced by neurotransmiters (presumably Dopaminergic -DA, Ventral Tegmental Area-VTA; Noradrenergic -NA, Locus Coeruleus-LC and Serotoninergic -5-HT, Raphe Nuclei- RN, respectively), (Otmakhova and Lisman, 1996; Katsuki et al., 1997), although the anatomical basis of the chemical modulation of memory in the HF is far from being understood. The neuroanatomical connections between the brainstem and in the HF in the nonhuman primate are still unclear. Previous tracer studies showed retrogradely labeled neurons in the brainstem areas including the VTA, LC and RN, after deposits in the hippocampus (Amaral and Cowan, 1980), as well as in the EC (Insausti et al., 1987). In order to characterize the neurochemical nature of those projections, as well as their topographic and laminar differences, we studied comparatively the distribution on those substances in the HF using immunohistochemical techniques. Immunohistochemistry for each DA (Tyrosine Hydroxylase, TH), NA (Dopamine Beta Hydroxylase -DBH-, and 5-HT) as well as double-immunohistochemical techniques using Alexa 488 (5-HT detection) and Alexa 568 (TH or DBH labeling) disclosed that: • The polymorphic layer of the DG had fibers with the three neurotransmitters, whereas the molecular layer showed only TH and 5-HT immunolabeling, without double-stained processes. • The pyramidal layer of CA3 showed denser 5-HT fiber labeling than TH; CA1 showed only scattered TH and 5-HT fibers, without double labeling profiles. • The subiculum and presubiculum showed fibers immunoreactive for TH, SER and BHD in the molecular layer. No double-labeled TH-5HT or DBH-5HT fibers were seen. • The superficial layers of the rostral EC (I and II) displayed TH- or 5-HT-labelled processes, while the most lateral subdivisions of EC (ELR/ELc) had TH- or DBH-positive fibers; they did not show co-localization. The preferential location of these positive fibers in ELR/ELc is significant, as this portion of the EC receives abundant unimodal and polymodal sensory input and innervates the body and tail of the hippocampus, and therefore it might be an important step for the monoaminergic modulation memory consolidation. Our preliminary anatomical results suggest that the HF function may be modulated independently by monoaminergic neurotransmitters

Frontal cortex afferents to the ventral tegmental area in the Macaca fascicularis monkey

The prefrontal cortico-midbrain pathway is thought to play an important role in the regulation of the firing pattern in the ventral tegmental area (VTA) neurons. The understanding of the mechanisms that underlie the regulation of the midbrain dopamine neurons is critical to elucidate the reward system as well as certain pathological conditions such as drug addiction or schizophrenia. Descending prefrontal cortex (PFC) projections to the VTA have been primarily documented in the rodent brain (Maurice et al., 1999; Sesack and Carr, 2002). Furthermore, several anatomical studies based on the use of anterograde tracers in the nonhuman primate, have shown labeled fibers in the VTA that originated in the medial frontal cortex and anterior cingulate cortex (areas 25, 32 and 24), orbitofrontal cortex (areas 11 and 14) and dorsolateral prefrontal cortex (area 9 and 46) (Chiba et al., 2001; Frankle et al., 2006). In order to complete the study of the direct inputs from the PFC to the VTA, the retrograde tracer 3 Fast Blue (FB) was placed in the mesencephalic ventral and dorsal tegmentum in Macaca fascicularis monkey, including the ventral tegmental area. We analyzed three cases injected with FB through a Hamilton syringe in the ventral mesencephalon. A magnetic resonance (MR) examination to localize the stereotaxic coordinates of the injection site was performed in all the animals used in this study. After 2 weeks survival, animals were deeply anesthetized and perfused through the heart with 4 paraformaldehyde. Several additional cases with 3H-aminoacid injections reported previously (Insausti and Amaral, 2008) were also available for analysis under dark field illumination. Our preliminary results showed labeled neurons in the deep layers of principally, the medial frontal and orbitofrontal cortices, including areas 24, 32 and 25, and the orbitofrontal cortex (areas 11, 13, 12 and 14). Comparatively, the dorsolateral prefrontal (area 10, 9, 46 and 6) cortex displayed far fewer labeled neurons. Most of the labeled neurons were situated at the level of the medial part of caudal area 9 and rostral area 6. The anterograde tracer experiments (5 cases with 3H-aminoacid deposits placed in the orbitofrontal cortex, and 3 cases in the medial frontal cortex) confirmed the existence of these projections, thus ruling out the contamination by fibers of passage at the retrograde tracer injection sites. Our data suggest that the influence of medial frontal and orbitofrontal cortices on the dopaminergic ascending projections is much higher than from the dorsolateral prefrontal cortex