VTA Excitatory Neurons Control Reward-driven Behavior by Modulating Infralimbic Cortical Firing.

The functional dichotomy of anatomical regions of the medial prefrontal cortex (mPFC) has been tested with greater certainty in punishment-driven tasks, and less so in reward-oriented paradigms. In the infralimbic cortex (IL), known for behavioral suppression (STOP), tasks linked with reward or punishment are encoded through firing rate decrease or increase, respectively. Although the ventral tegmental area (VTA) is the brain region governing reward/aversion learning, the link between its excitatory neuron population and IL encoding of reward-linked behavioral expression is unclear. Here, we present evidence that IL ensembles use a population-based mechanism involving broad inhibition of principal cells at intervals when reward is presented or expected. The IL encoding mechanism was consistent across multiple sessions with randomized rewarded target sites. Most IL neurons exhibit FR (Firing Rate) suppression during reward acquisition intervals (T1), and subsequent exploration of previously rewarded targets when the reward is omitted (T2). Furthermore, FR suppression in putative IL ensembles persisted for intervals that followed reward-linked target events. Pairing VTA glutamate inhibition with reward acquisition events reduced the weight of reward-target association expressed as a lower affinity for previously rewarded targets. For these intervals, fewer IL neurons per mouse trial showed FR decrease and were accompanied by an increase in the percentage of units with no change in FR. Together, we conclude that VTA glutamate neurons are likely involved in establishing IL inhibition states that encode reward acquisition, and subsequent reward-target association.

VTA multifaceted modulation of CA1 local circuits.

Excitatory pyramidal (PYR) cell activation of interneurons (INT) produces network oscillations that underlie cognitive processes in the hippocampus (CA1). Neural projections from the ventral tegmental area (VTA) to the hippocampus contribute to novelty detection by modulating CA1 PYR and INT activity. The role of the VTA in the VTA-hippocampus loop is mostly attributed to the dopamine neurons although the VTA glutamate-releasing terminals are dominant in the hippocampus. Because of the traditional focus on VTA dopamine circuits, how VTA glutamate inputs modulate PYR activation of INT in CA1 neuronal ensembles is poorly understood and has not been distinguished from the VTA dopamine inputs. By combining CA1 extracellular recording with VTA photostimulation in anesthetized mice, we compared the effects of VTA dopamine and glutamate input on CA1 PYR/INT connections. Stimulation of VTA glutamate neurons shortened PYR/INT connection time without altering the synchronization or connectivity strength. Conversely, activation of VTA dopamine inputs delayed CA1 PYR/INT connection time and increased the synchronization in putative pairs. Taken together, we conclude that VTA dopamine and glutamate projections produce tract-specific effects on CA1 PYR/INT connectivity and synchrony. As such, selective activation or co-activation of these systems will likely produce a range of modulatory effects on local CA1 circuits.

Anterior thalamic glutamatergic neurons modulate prefrontal cortical firing and regularity.

For decades, studies on learning and memory focused extensively on the neural circuitry between the hippocampus and the prefrontal cortex, with minimal consideration of the role of the anterior thalamic nucleus (ATN). The diencephalic nucleus is rich in excitatory glutamate neurons that project to both the hippocampus and medial prefrontal cortex (mPFC). Neural projections from the hippocampus and mPFC have also been identified in the ATN through reciprocal circuits. Although ATN lesioning leads to memory loss (amnesia), the role of the ATN in the propagation of cognitive processes in the mPFC is still poorly understood. In the current study, we employed adeno-associated viral labeling and in vivo electrophysiology to trace ATN glutamate neural projections in the layers of the mPFC. Neuroanatomical mapping of ATN Vglut2+ projections revealed a topographic gradient in the infralimbic cortex (IL), prelimbic cortex (PrL), and cingulate cortex (Cg1). Specifically, the IL has the least ATN Vglut2+ terminals while the PrL (layer III) and Cg1 (layers III, Va/b, VI) have robust innervation. Functional tracing of the thalamocortical projection was performed by combining extracellular Cg1 and ATN recording with ATN Vglut2+ neuron photostimulation. Our results show that light-activated ATN Vglut2+ neurons drive Cg1 neural activity and increase the firing rate of putative pyramidal neurons in anesthetized mice. In addition, Cg1 putative neurons that were activated during ATN photostimulation show a significant increase in firing regularity in comparison with the baseline (no ATN stimulus). Together, our results show that ATN Vglut2+ neurons modulate the firing rate and regularity of the putative Cg1 pyramidal cells, thus, creating a premise for direct influence on cognitive processes in cortical networks.

CA1 Spike Timing is Impaired in the 129S Inbred Strain During Cognitive Tasks.

A spontaneous mutation of the disrupted in schizophrenia 1 (Disc1) gene is carried by the 129S inbred mouse strain. Truncated DISC1 protein in 129S mouse synapses impairs the scaffolding of excitatory postsynaptic receptors and leads to progressive spine dysgenesis. In contrast, C57BL/6 inbred mice carry the wild-type Disc1 gene and exhibit more typical cognitive performance in spatial exploration and executive behavioral tests. Because of the innate Disc1 mutation, adult 129S inbred mice exhibit the behavioral phenotypes of outbred B6 Disc1 knockdown (Disc1-/-) or Disc1-L-100P mutant strains. Recent studies in Disc1-/- and L-100P mice have shown that impaired excitation-driven interneuron activity and low hippocampal theta power underlie the behavioral phenotypes that resemble human depression and schizophrenia. The current study compared the firing rate and connectivity profile of putative neurons in the CA1 of freely behaving inbred 129S and B6 mice, which have mutant and wild-type Disc1 genes, respectively. In cognitive behavioral tests, 129S mice had lower exploration scores than B6 mice. Furthermore, the mean firing rate for 129S putative pyramidal (pyr) cells and interneurons (int) was significantly lower than that for B6 CA1 neurons sampled during similar tasks. Analysis of pyr/int connectivity revealed a significant delay in synaptic transmission for 129S putative pairs. Sampled 129S pyr/int pairs also had lower detectability index scores than B6 putative pairs. Therefore, the spontaneous Disc1 mutation in the 129S strain attenuates the firing of putative pyr CA1 neurons and impairs spike timing fidelity during cognitive tasks.

Combined In Vivo Anatomical and Functional Tracing of Ventral Tegmental Area Glutamate Terminals in the Hippocampus.

Optogenetic modulation of neuron sub-populations in the brain has allowed researchers to dissect neural circuits in vivo and ex vivo. This provides a premise for determining the role of neuron types within a neural circuit, and their significance in information encoding relative to learning. Likewise, the method can be used to test the physiological significance of two or more connected brain regions in awake and anesthetized animals. The current study demonstrates how VTA glutamate neurons modulate the firing rate of putative pyramidal neurons in the CA1 (hippocampus) of anesthetized mice. This protocol employs adeno-associated virus (AAV)-dependent labeling of VTA glutamate neurons for the tracing of VTA presynaptic glutamate terminals in the layers of the hippocampus. Expression of light-controlled opsin (channelrhodopsin; hChR2) and fluorescence protein (eYFP) harbored by the AAV vector permitted anterograde tracing of VTA glutamate terminals, and photostimulation of VTA glutamate neuron cell bodies (in the VTA). High-impedance acute silicon electrodes were positioned in the CA1 to detect multi-unit and single-unit responses to VTA photostimulation in vivo. The results of this study demonstrate the layer-dependent distribution of presynaptic VTA glutamate terminals in the hippocampus (CA1, CA3, and DG). Also, the photostimulation of VTA glutamate neurons increased the firing and burst rate of putative CA1 pyramidal units in vivo.

Distribution of VTA Glutamate and Dopamine Terminals, and their Significance in CA1 Neural Network Activity.

Reciprocal connection between the ventral tegmental area (VTA) and the hippocampus forms a loop that controls information entry into long-term memory. Compared with the widely studied VTA dopamine system, VTA glutamate terminals are anatomically dominant in the hippocampus and less understood. The current study employs anterograde and retrograde labeling of VTA dopamine and glutamate neurons to map the distribution of their terminals within the layers of the hippocampus. Also, functional tracing of VTA dopamine and glutamate projections to the hippocampus was performed by photostimulation of VTA cell bodies during CA1 extracellular voltage sampling in vivo. VTA dopamine terminals predominantly innervate the CA1 basal dendrite layer and modulate the firing rate of active putative neurons. In contrast, anatomical dominance of VTA glutamate terminals in the CA1 pyramidal cell and apical dendrite layers suggests the possible involvement of these terminals in excitability regulation. In support of these outcomes, photostimulation of VTA dopamine neurons increased the firing rate but not intrinsic excitability parameters for putative pyramidal units. Conversely, activation of VTA glutamate neurons increased CA1 network firing rate and burst rate. In addition, VTA glutamate inputs reduced the interspike and interburst intervals for putative CA1 neurons. Taken together, we deduced that layer-specific distribution of presynaptic dopamine and glutamate terminals in the hippocampus determinines VTA modulation (dopamine) or regulation (glutamate) of excitability in the CA1 neural network.

Positive Modulation of SK Channel Impedes Neuron-Specific Cytoskeletal Organization and Maturation.

N-methyl-D-aspartate receptor (NMDAR) modulates the structural plasticity of dendritic spines by impacting cytoskeletal organization and kinase signaling. In the developing nervous system, activation of NMDAR is pertinent for neuronal migration, neurite differentiation, and cellular organization. Given that small conductance potassium channels (SK2/3) repress NMDAR ionotropic signaling, this study highlights the impact of neonatal SK channel potentiation on adult cortical and hippocampal organization. Neonatal SK channel potentiation was performed by one injection of SK2/3 agonist (CyPPA) into the pallium of mice on postnatal day 2 (P2). When the animals reached adulthood (P55), the hippocampus and cortex were examined to assess neuronal maturation, lamination, and the distribution of synaptic cytoskeletal proteins. Immunodetection of neuronal markers in the brain of P2-treated P55 mice revealed the presence of immature neurons in the upper cortical layers (layers II-IV) and CA1 (hippocampus). Also, layer-dependent cortical-cell density was attenuated due to the ectopic localization of mature (NeuN+) and immature (Doublecortin+ [DCX+]) neurons in cortical layers II-IV. Similarly, the decreased count of NeuN+ neurons in the CA1 is accompanied by an increase in the number of immature DCX+ neurons. Ectopic localization of neurons in the upper cortex and CA1 caused the dramatic expression of neuron-specific cytoskeletal proteins. In line with this, structural deformity of neuronal projections and the loss of postsynaptic densities suggests that postsynaptic integrity is compromised in the SK2/3+ brain. From these results, we deduced that SK channel activity in the developing brain likely impacts neuronal maturation through its effects on cytoskeletal formation.

Disc1 Carrier Mice Exhibit Alterations in Neural pIGF-1Rß and Related Kinase Expression.

Mutation of the disc1 gene underlies a broad range of developmental neuropsychiatric defects, including schizophrenia, depression, and bipolar disorder. The pathophysiological phenotypes linked with disc1 mutation are due to the truncation of the DISC1 primary protein structure. This leads to a defective post-synaptic scaffolding and kinase-GSK3ß and Erk1/2-signaling. As a result, synaptic function and maintenance are significantly impaired in the disc1 mutant brain. Among several other pathways, GSK3ß and Erk1/2 are involved in insulin-like growth factor 1 receptor (IGF-1Rß) kinase signaling. Although disc1 mutation alters these kinases, it is unclear if the mutation impacts IGF-1R expression and activity in the brain. Here, we demonstrate that the expression of active IGF-1Rß (pIGF-1Rß) is altered in the hippocampus and prefrontal cortex (PFC) of disc1 mutant mice and vary with the dose of the mutation (homozygous and heterozygous). The expression of pIGF-1Rß decreased significantly in 129S (hom, disc1 -/-) brains. In contrast, 129S:B6 (het, disc1 +/-) brains were characterized by an increase in pIGF-1Rß when compared with the C57BL/6 (disc1 +/+) level. The decrease in pIGF-1Rß level for the 129S brains was accompanied by the loss of Akt activity (S473 pAkt) and decreased Ser9 phosphorylation of GSK3ß (increased basal GSK3ß). Additionally, hippocampal and cortical pErk1/2 activity increased in the 129S hippocampus and cortex. Although 129S:B6 recorded alterations in pIGF-1Rß-pAkt-GSK3ß (like 129S), there was no observable change in pErk1/2 activity for the heterozygote (disc1 +/-) mutant. In addition to GSK3ß inhibition, we conclude that pIGF-1R, pAkt, and pErk1/2 are potential targets in disc1 -/- mutant brain. On the other hand, pIGF-1R and pAkt can be further explored in disc1 +/- brain.

SK Channel Modulates Synaptic Plasticity by Tuning CaMKIIα/ß Dynamics.

N-Methyl-D-Aspartate Receptor 1 (NMDAR)-linked Ca++ current represents a significant percentage of post-synaptic transient that modulates synaptic strength and is pertinent to dendritic spine plasticity. In the hippocampus, Ca++ transient produced by glutamatergic ionotropic neurotransmission facilitates Ca++-Calmodulin-dependent kinase 2 (CaMKII) Thr286 phosphorylation and promote long-term potentiation (LTP) expression. At CA1 post-synaptic densities, Ca++ transients equally activate small conductance (SK2) channel which regulates excitability by suppressing Ca++ movement. Here, we demonstrate that upstream attenuation of GluN1 function in the hippocampus led to a decrease in Thr286 CaMKIIα phosphorylation, and increased SK2 expression. Consistent with the loss of GluN1 function, potentiation of SK channel in wild type hippocampus reduced CaMKIIα expression and abrogate synaptic localization of T286 pCaMKIIα. Our results demonstrate that positive modulation of SK channel at hippocampal synapses likely refine GluN1-linked plasticity by tuning dendritic localization of CaMKIIα.

Contrasting characteristic behaviours among common laboratory mouse strains.

Mice are widely used to model wide-ranging human neurological disorders, from development to degenerative pathophysiology. Behavioural and molecular characteristics of these mouse models are influenced by the genetic background of each strain. Among the most commonly used strains, the inbred C57BL/6J, BALB/c, CBA and 129SvEv lines and the CD1 outbred line are particularly predominant. Despite their prevalence, comparative performance of these strains on many standard behavioural tests commonly used to assess neurological conditions remains diffusely and indirectly accessible in the literature. Given that independent studies may be conducted with mice of differing genetic backgrounds, any variation in characteristic behavioural responses of specific strains should be delineated in order to properly interpret results among studies. Thus, in the present study, we aimed to characterize these commonly used mice strains through several standard behavioural tests. Here, we found that animals from different genetic background strains exhibited varying behavioural patterns when assessed for sociability/novelty, memory function, and negative behaviours like despair and stress calls. These results suggest that genetic variation among strains may be responsible-in part-for strain-specific behavioural phenotypes and potential predisposition to some neurological disorders.

A Putative Mechanism of Age-Related Synaptic Dysfunction Based on the Impact of IGF-1 Receptor Signaling on Synaptic CaMKIIα Phosphorylation.

Insulin-like growth factor 1 receptor (IGF-1R) signaling regulates the activity and phosphorylation of downstream kinases linked to inflammation, neurodevelopment, aging and synaptic function. In addition to the control of Ca2+ currents, IGF-1R signaling modulates the activity of calcium-calmodulin-dependent kinase 2 alpha (CaMKIIα) and mitogen activated protein kinase (MAPK/ErK) through multiple signaling pathways. These proteins (CaMKIIα and MAPK) regulate Ca2+ movement and long-term potentiation (LTP). Since IGF-1R controls the synaptic activity of Ca2+, CaMKIIα and MAPK signaling, the possible mechanism through which an age-dependent change in IGF-1R can alter the synaptic expression and phosphorylation of these proteins in aging needs to be investigated. In this study, we evaluated the relationship between an age-dependent change in brain IGF-1R and phosphorylation of CaMKIIα/MAPK. Furthermore, we elucidated possible mechanisms through which dysregulated CaMKIIα/MAPK interaction may be linked to a change in neurotransmitter processing and synaptic function. Male C57BL/6 VGAT-Venus mice at postnatal days 80 (P80), 365 and 730 were used to study age-related neural changes in two brain regions associated with cognitive function: hippocampus and prefrontal cortex (PFC). By means of high throughput confocal imaging and quantitative immunoblotting, we evaluated the distribution and expression of IGF-1, IGF-1R, CaMKIIα, p-CaMKIIα, MAPK and p-MAPK in whole brain lysate, hippocampus and cortex. Furthermore, we compared protein expression patterns and regional changes at P80, P365 and P730. Ultimately, we determined the relative phosphorylation pattern of CaMKIIα and MAPK through quantification of neural p-CaMKIIα and p-MAPK/ErK, and IGF-1R expression for P80, P365 and P730 brain samples. In addition to a change in synaptic function, our results show a decrease in neural IGF-1/IGF-1R expression in whole brain, hippocampus and cortex of aged mice. This was associated with a significant upregulation of phosphorylated neural MAPK (p-MAPK) and decrease in total brain CaMKIIα (i.e., CaMKIIα and p-CaMKIIα) in the aged brain. Taken together, we showed that brain aging is associated with a change in neural IGF-1/IGF-1R expression and may be linked to a change in phosphorylation of synaptic kinases (CaMKIIα and MAPK) that are involved in the modulation of LTP.

IGF-I Gene Therapy in Aging Rats Modulates Hippocampal Genes Relevant to Memory Function.

In rats, learning and memory performance decline during normal aging, which makes this rodent species a suitable model to evaluate therapeutic strategies. In aging rats, insulin-like growth factor-I (IGF-I), is known to significantly improve spatial memory accuracy as compared to control counterparts. A constellation of gene expression changes underlie the hippocampal phenotype of aging but no studies on the effects of IGF-I on the hippocampal transcriptome of old rodents have been documented. Here, we assessed the effects of IGF-I gene therapy on spatial memory performance in old female rats and compared them with changes in the hippocampal transcriptome. In the Barnes maze test, experimental rats showed a significantly higher exploratory frequency of the goal hole than controls. Hippocampal RNA-sequencing showed that 219 genes are differentially expressed in 28-month-old rats intracerebroventricularly injected with an adenovector expressing rat IGF-I as compared with placebo adenovector-injected counterparts. From the differentially expressed genes, 81 were down and 138 upregulated. From those genes, a list of functionally relevant genes, concerning hippocampal IGF-I expression, synaptic plasticity as well as neuronal function was identified. Our results provide an initial glimpse at the molecular mechanisms underlying the neuroprotective actions of IGF-I in the aging brain.

Systemic Sympathoexcitation Was Associated with Paraventricular Hypothalamic Phosphorylation of Synaptic CaMKIIα and MAPK/ErK.

Systemic administration of adrenergic agonist (Isoproterenol; ISOP) is known to facilitate cardiovascular changes associated with heart failure through an upregulation of cardiac toll-like receptor 4 (TLR4). Furthermore, previous studies have shown that cardiac tissue-specific deletion of TLR4 protects the heart against such damage. Since the autonomic regulation of systemic cardiovascular function originates from pre-autonomic sympathetic centers in the brain, it is unclear how a systemically driven sympathetic change may affect the pre-autonomic paraventricular hypothalamic nuclei (PVN) TLR4 expression. Here, we examined how change in PVN TLR4 was associated with alterations in the neurochemical cytoarchitecture of the PVN in systemic adrenergic activation. After 48 h of intraperitoneal 150 mg/kg ISOP treatment, there was a change in PVN CaMKIIα and MAPK/ErK expression, and an increase in TLR4 in expression. This was seen as an increase in p-MAPK/ErK, and a decrease in synaptic CaMKIIα expression in the PVN (p < 0.01) of ISOP treated mice. Furthermore, there was an upregulation of vesicular glutamate transporter (VGLUT 2; p < 0.01) and a decreased expression of GABA in the PVN of Isoproterenol (ISOP) treated WT mice (p < 0.01). However, after a PVN-specific knockdown of TLR4, the effect of systemic administration of ISOP was attenuated, as indicated by a decrease in p-MAPK/ErK (p < 0.01) and upregulation of CaMKIIα (p < 0.05). Additionally, loss of inhibitory function was averted while VGLUT2 expression decreased when compared with the ISOP treated wild type mice and the control. Taken together, the outcome of this study showed that systemic adrenergic activation may alter the expression, and phosphorylation of preautonomic MAPK/ErK and CaMKIIα downstream of TLR4. As such, by outlining the roles of these kinases in synaptic function, we have identified the significance of neural TLR4 in the progression, and attenuation of synaptic changes in the pre-autonomic sympathetic centers.

Stress-altered synaptic plasticity and DAMP signaling in the hippocampus-PFC axis; elucidating the significance of IGF-1/IGF-1R/CaMKIIα expression in neural changes associated with a prolonged exposure therapy.

Traumatic stress patients showed significant improvement in behavior after a prolonged exposure to an unrelated stimulus. This treatment method attempts to promote extinction of the fear memory associated with the initial traumatic experience. However, the subsequent prolonged exposure to such stimulus creates an additional layer of neural stress. Although the mechanism remains unclear, prolonged exposure therapy (PET) likely involves changes in synaptic plasticity, neurotransmitter function and inflammation; especially in parts of the brain concerned with the formation and retrieval of fear memory (Hippocampus and Prefrontal Cortex: PFC). Since certain synaptic proteins are also involved in danger-associated molecular pattern signaling (DAMP), we identified the significance of IGF-1/IGF-1R/CaMKIIα expression as a potential link between the concurrent progression of synaptic and inflammatory changes in stress. Thus, a comparison between IGF-1/IGF-1R/CaMKIIα, synaptic and DAMP proteins in stress and PET may highlight the significance of PET on synaptic morphology and neuronal inflammatory response. In behaviorally characterized Sprague-Dawley rats, there was a significant decline in neural IGF-1 (p<0.001), hippocampal (p<0.001) and cortical (p<0.05) IGF-1R expression. These animals showed a significant loss of presynaptic markers (synaptophysin; p<0.001), and changes in neurotransmitters (VGLUT2, Tyrosine hydroxylase, GABA, ChAT). Furthermore, naïve stressed rats recorded a significant decrease in post-synaptic marker (PSD-95; p<0.01) and synaptic regulator (CaMKIIα; p<0.001). As part of the synaptic response to a decrease in brain CaMKIIα, small ion conductance channel (KCa2.2) was upregulated in the brain of naïve stressed rats (p<0.01). After a PET, an increase in IGF-1 (p<0.05) and IGF-1R was recorded in the Stress-PET group (p<0.001). As such, hippocampal (p<0.001), but not cortical (ns) synaptophysin expression increased in Stress-PET. Although PSD-95 was relatively unchanged in the hippocampus and PFC, CaMKIIα (p<0.001) and KCa2.2 (p<0.01) were upregulated in Stress-PET, and may be involved in extinction of fear memory-related synaptic potentials. These changes were also associated with a normalized neurotransmitter function, and a significant reduction in open space avoidance; when the animals were assessed in elevated plus maze (EPM). In addition to a decrease in IGF-1/IGF-1R, an increase in activated hippocampal and cortical microglia was seen in stress (p<0.05) and after a PET (Stress-PET; p<0.001). Furthermore, this was linked with a significant increase in HMGB1 (Hippocampus: p<0.001, PFC: p<0.05) and TLR4 expression (Hippocampus: p<0.01; PFC: ns) in the neurons. Taken together, this study showed that traumatic stress and subsequent PET involves an event-dependent alteration of IGF1/IGF-1R/CaMKIIα. Firstly, we showed a direct relationship between IGF-1/IGF-1R expression, presynaptic function (synaptophysin) and neurotransmitter activity in stress and PET. Secondly, we identified the possible role of CaMKIIα in post-synaptic function and regulation of small ion conductance channels. Lastly, we highlighted some of the possible links between IGF1/IGF-1R/CaMKIIα, the expression of DAMP proteins, Microglia activation, and its implication on synaptic plasticity during stress and PET.

Long-lasting training in the Barnes maze prompts hippocampal spinogenesis and habituation in rats.

There is a constant need to assess spatial memory in small rodents to elucidate the basics of cognition in neuroscience experiments. Thus, the significance of the Barnes maze in the biology of hippocampal and cortical neural function cannot be overemphasized. Despite the wide use of the Barnes maze, the effect of maze task training on the structure of hippocampal neurons is yet to be elucidated. Adult Sprague-Dawley rats were subjected to intense training on the Barnes maze (3 months). Subsequently, the hippocampus (cornus ammonis and dentate gyrus) of separate sets of rats was processed for Golgi Colonnier techniques (silver impregnation) and adenoviral-green fluorescent protein labeling (immunohistochemistry). Our results showed that training the animals on the Barne maze increased spinogenesis significantly in the cornus ammonis and dentate gyrus neurons. In addition, we identified a critical time point at which the rats habituated to the trial without escaping box (the probe trial) and could not be tested further in the maze. Taken together, we deduced that a prolonged test on the dry land maze facilitated habituation and caused an increase in hippocampal dendritic spine count. As such, the dry land maze is a suitable paradigm for assessing spatial memory in rats. However, precautions should be taken in selecting suitable experimental controls on the basis of the duration of a study.

Thalamic dopaminergic neurons projects to the paraventricular nucleus-rostral ventrolateral medulla/C1 neural circuit.

Paraventricular nuclei (PVN) projections to the rostral ventrolateral medulla (RVLM)/C1 catecholaminergic neuron group constitute the pre-autonomic sympathetic center involved in the neural control of systemic cardiovascular function. However, the role of extra-hypothalamic and thalamic dopaminergic (DA) inputs in this circuit remains underexplored. Using retrograde neuroanatomical tracing and high contrast confocal imaging methods, we investigated the projections and morphology of the discrete thalamic DA neuron groups in the dorsal hypothalamic area and their contribution to the PVN-RVLM neural circuit. We found that DA neuron subgroups in the Zona Incerta (Zi; 60%) and Reuniens thalamic nuclei (Re; 40%) were labeled comparably to the PVN (85%) after a retrograde tracer was injected into the RVLM/C1 (P < 0.01 mean ± SEM). The Re/Zi DA neuron subgroups were characterized by angulated cell bodies, superiomedial and inferiomedial projections reaching the contralateral Re/Zi and ipsilateral PVN DA neurons respectively. Ultimately, we deduced that the DA projections of the Re/Zi to the PVN contribute to the PVN-RVLM/C1 neural circuit. As a result of these connections, the Re/Zi DA neuron groups may regulate preautonomic sympathetic events associated with the PVN-RVLM pathway. Anat Rec, 300:1307-1314, 2017. © 2016 Wiley Periodicals, Inc.

Age-dependent alterations to paraventricular nucleus insulin-like growth factor 1 receptor as a possible link between sympathoexcitation and inflammation.

Modifications to neural circuits of the paraventricular hypothalamic nucleus (PVN) have been implicated in sympathoexcitation and systemic cardiovascular dysfunction. However, to date, the role of insulin-like growth factor 1 receptor (IGF-1R) expression on PVN pathophysiology is unknown. Using confocal immunofluorescence quantification and electrophysiological recordings from acute PVN slices, we investigated the mechanism through which age-dependent IGF-1R depletion contributes to the progression of inflammation and sympathoexcitation in the PVN of spontaneously hypertensive rats (SHR). Four and twenty weeks old SHR and Wistar Kyoto (WKY) rats were used for this study. Our data showed that angiotensin I/II and pro-inflammatory high mobility box group protein 1 (HMGB1) exhibited increased expression in the PVN of SHR versus WKY at 4 weeks (p < 0.01), and were even more highly expressed with age in SHR (p < 0.001). This correlated with a significant decrease in IGF-1R expression, with age, in the PVN of SHR when compared with WKY (p < 0.001) and were accompanied by related changes in astrocytes and microglia. In subsequent analyses, we found an age-dependent change in the expression of proteins associated with IGF-1R signaling pathways involved in inflammatory responses and synaptic function in the PVN. MAPK/ErK was more highly expressed in the PVN of SHR by the fourth week (p < 0.001; vs. WKY), while expression of neuronal nitric oxide synthase (p < 0.001) and calcium-calmodulin-dependent kinase II alpha (CamKIIα; p < 0.001) were significantly decreased by the 4th and 20th week, respectively. Age-dependent changes in MAPK/ErK expression in the PVN correlated with an increase in the expression of vesicular glutamate transporter (p < 0.001 vs. WKY), while decreased levels of CamKIIα was associated with a decreased expression of tyrosine hydroxylase (p < 0.001) by the 20th week. In addition, reduced labeling for ϒ-aminobutyric acid in the PVN of SHR (p < 0.001) correlated with a decrease in neuronal nitric oxide synthase labeling (p < 0.001) when compared with the WKY by the 20th week. Electrophysiological recordings from neurons in acute slice preparations of the PVN of 4 weeks old SHR revealed spontaneous post-synaptic currents of higher frequency when compared with neurons from WKY PNV slices of the same age (p < 0.001; n = 14 cells). This also correlated with an increase in PSD-95 in the PVN of SHR when compared with the WKY (p < 0.001). Overall, we found an age-dependent reduction of IGF-1R, and related altered expression of associated downstream signaling molecules that may represent a link between the concurrent progression of synaptic dysfunction and inflammation in the PVN of SHR.

Neural and behavioural changes in male periadolescent mice after prolonged nicotine-MDMA treatment.

The interaction between MDMA and Nicotine affects multiple brain centres and neurotransmitter systems (serotonin, dopamine and glutamate) involved in motor coordination and cognition. In this study, we have elucidated the effect of prolonged (10 days) MDMA, Nicotine and a combined Nicotine-MDMA treatment on motor-cognitive neural functions. In addition, we have shown the correlation between the observed behavioural change and neural structural changes induced by these treatments in BALB/c mice. We observed that MDMA (2 mg/Kg body weight; subcutaneous) induced a decline in motor function, while Nicotine (2 mg/Kg body weight; subcutaneous) improved motor function in male periadolescent mice. In combined treatment, Nicotine reduced the motor function decline observed in MDMA treatment, thus no significant change in motor function for the combined treatment versus the control. Nicotine or MDMA treatment reduced memory function and altered hippocampal structure. Similarly, a combined Nicotine-MDMA treatment reduced memory function when compared with the control. Ultimately, the metabolic and structural changes in these neural systems were seen to vary for the various forms of treatment. It is noteworthy to mention that a combined treatment increased the rate of lipid peroxidation in brain tissue.

Kolaviron was protective against sodium azide (NaN3) induced oxidative stress in the prefrontal cortex.

Kolaviron is a phytochemical isolated from Garcina kola (G. kola); a common oral masticatory agent in Nigeria (West Africa). It is a bioflavonoid used--as an antiviral, anti-inflammatory and antioxidant--in relieving the symptoms of several diseases and infections. In this study we have evaluated the neuroprotective and regenerative effect of kolaviron in neurons of the prefrontal cortex (Pfc) before or after exposure to sodium azide (NaN3) induced oxidative stress. Separate groups of animals were treated as follows; kolaviron (200 mg/Kg) for 21 days; kolaviron (200 mg/Kg for 21 days) followed by NaN3 treatment (20 mg/Kg for 5 days); NaN3 treatment (20 mg/Kg for 5 days) followed by kolaviron (200 mg/Kg for 21 days); 1 ml of corn-oil (21 days-vehicle); NaN3 treatment (20 mg/Kg for 5 days). Exploratory activity associated with Pfc function was assessed in the open field test (OFT) following which the microscopic anatomy of the prefrontal cortex was examined in histology (Haematoxylin and Eosin) and antigen retrieval Immunohistochemistry to show astroglia activation (GFAP), neuronal metabolism (NSE), cytoskeleton (NF) and cell cycle dysregulation (p53). Subsequently, we quantified the level of Glucose-6-phosphate dehydrogenase (G6PDH) and lactate dehydrogenase (LDH) in the brain tissue homogenate as a measure of stress-related glucose metabolism. Kolaviron (Kv) and Kolaviron/NaN3 treatment caused no prominent change in astroglia density and size while NaN3 and NaN3/Kv induced astroglia activation and scar formation (astrogliosis) in the Pfc when compared with the control. Similarly, Kolaviron and Kv/NaN3 did not alter NSE expression (glucose metabolism) while NaN3 and NaN3/Kv treatment increased cortical NSE expression; thus indicating stress related metabolism. Further studies on enzymes of glucose metabolism (G6PDH and LDH) showed that NaN3 increased LDH while kolaviron reduced LDH in the brain tissue homogenate (P < 0.001). In addition kolaviron treatment before (P < 0.001) or after (P < 0.05) NaN3 treatment also reduced LDH expression; thus supporting its role in suppression of oxidative stress. Interestingly, NF deposition increased in the Pfc after kolaviron treatment while Kv/NaN3 showed no significant change in NF when compared with the control. In furtherance, NaN3 and NaN3/Kv caused a decrease in NF deposition (degeneration). Ultimately, the protective effect of KV administered prior to NaN3 treatment was confirmed through p53 expression; which was similar to the control. However, NaN3 and NaN3/Kv caused an increase in p53 expression in the Pfc neurons (cell cycle dysregulation). We conclude that kolaviron is not neurotoxic when used at 200 mg/Kg BW. Furthermore, 200 mg/Kg of kolaviron administered prior to NaN3 treatment (Kv/NaN3) was neuroprotective when compared with Kolaviron administered after NaN3 treatment (NaN3/Kv). Some of the observed effects of kolaviron administered before NaN3 treatment includes reduction of astroglia activation, absence of astroglia scars, antioxidation (reduced NSE and LDH), prevention of neurofilament loss and cell cycle regulation.