Primary sensory neurons deliver information from the periphery to specific circuits in the central nervous system. It is vital that each sensory neuron detects the appropriate type of stimulus and conveys that information to appropriate regions of the sensory neuropil to target second-order neurons. Molecular programs that coordinate sensory morphology in the periphery with axon projection patterns centrally are poorly understood. I have used the multidendritic (md) sensory neurons of the Drosophila melanogaster peripheral nervous system to identify genetic and molecular programs that coordinate dendrite and axonal morphogenesis in individual sensory neurons. The homeodomain transcription factor Cut is expressed in neurons with complex dendrite morphologies that innervate the epidermis and ventral axon projections in the CNS, and is absent from putative proprioceptive neurons that have simpler dendrites and target to more dorsal CNS regions. In this thesis I demonstrate that, in defined subsets of sensory neurons, loss of Cut leads to dendritic transformation to a proprioceptive-type arbor that is accompanied by a dorsal shift in the termination of their axons in the CNS. Mechanistically, I show that Cut functions at least in part by repressing the expression of the POU domain transcription factors Pdm1 and Pdm2 (Pdm1/2), which are normally expressed only in proprioceptive neurons. Gain and loss of function studies further suggest instructive roles for Pdm1/2 in the development of proprioceptive dendritic arborization and axonal targeting. Together these results identify a transcriptional program that coordinately specifies proprioceptive dendrite morphology and sensory axon targeting to modality-specific domains of the CNS. Using a candidate based approached I have identified three molecular regulators of proprioceptive neuron dendrite morphology. In addition, gene profiling of sensory neurons forced to express Pdm2 has identified over 600 genes that show changes in expression when Pdm2 is misexpressed and that may mediate the effects of Pdm1/2 in directing proprioceptive dendrite and axon development. These profiling experiments pave the way for the identification of novel regulators of dendrite and axon morphogenesis that link transcriptional programs to specific morphologies with consequences for sensory circuit function.

Neurons develop highly stereotypic dendritic arbors that influence establishment of proper connections and integration of information they receive to generate an appropriate output. Morphogenesis of dendrites is coordinated by both cell-intrinsic and extrinsic factors. Recent studies have begun to elucidate how interactions between neurons shape dendrite morphogenesis. However, influence of the substrate upon which neurons grow their dendritic arbors in this process is relatively poorly understood. Here I have used the peripheral sensory neurons of the Drosophila larva that grow dendrites over epithelial cell substrates to gain insights into how interactions with the substrate may influence dendrite development. In this thesis, I present data showing that Basigin, an immunoglobulin superfamily member, has somatodendritic and axonal localization in sensory neurons, and is enriched at cell borders and beneath class IV dendrites in epithelial cells. Loss of function analyses indicate that Basigin is required both in neurons and epithelial cell substrates for proper morphogenesis of the highly complex dendrites of class IV sensory neurons. Reduced innervation of the dendritic field of basigin mutant neurons was observed even at an immature stage, indicating a requirement of Basigin in these neurons for developmental elaboration of dendritic arbors. Structure-function analysis revealed that membrane-tethering of Basigin on the neuronal surface is essential for its function. In addition, a highly conserved tri-basic motif consisting of positively charged residues that may bind cytoskeletal adaptor proteins is required for its function in neurons. Results of genetic interaction analysis suggest that Basigin-mediated regulation of dendrite morphogenesis does not involve Integrin and matrix metalloproteinases, both of which have been implicated in Basigin function in other cellular contexts. I show that Basigin exhibits genetic interaction with Tropomodulin, an actin-capping protein, suggesting that they function in the same molecular pathway in regulating dendrite development. Taken together, data presented in this thesis support a model in which interaction between Basigin on the surfaces of neurons and epithelial cells regulate the underlying cytoskeleton within dendrites to influence their development. Thus, these results identify a novel molecular pathway that may mediate communication between neurons and their substrates that is essential for proper dendrite morphogenesis.

The thalamus is a potent driver of cortical activity, even though cortical synapses onto layer 4 (L4) neurons outnumber thalamic synapses ten to one. Previous in vitro studies have suggested that enhanced efficacy of thalamocortical (TC) relative to corticocortical (CC) synapses explains the effectiveness of the thalamus. We investigated possible key anatomical and physiological differences between these inputs onto excitatory L4 neurons in vivo. We developed a high-throughput light microscopy method, validated by electron microscopy, to completely map the locations of synapses across an entire dendritic tree. This demonstrated that TC synapses are slightly more proximal to the soma than CC synapses, but detailed compartmental modeling predicted that dendritic filtering does not appreciably favor one synaptic class over another. Measurements of synaptic strength in intact animals revealed that both TC and CC synapses are weak and approximately equivalent. We conclude that thalamic potency relies, not on enhanced TC strength, but on coincident activation of converging inputs.

Working memory is a type of memory that is active only for a short period of time (Fuster and Alexander, 1971; Goldman-Rakic, 1992). A common example of working memory is our ability to hold a phone number in our minds transiently, until it is dialed. Working memory is critical for many cognitive tasks, such as making decisions and guiding subsequent actions (Goldman-Rakic, 1992; Wickelgren, 2001). Deficits in working memory are associated with numerous pathological conditions, including schizophrenia, attention deficit hyperactivity disorder, aging, and stress (Birnbaum et al., 2004; Goldman-Rakic, 1992; Goldman-Rakic and Selemon, 1997). Therefore, it is important to understand the neural basis of working memory. During performance of a working memory task, pyramidal neurons in prefrontal cortex (PFC) are able to maintain sustained firing during a delay period between an informative cue and the appropriate behavioral response (Goldman-Rakic, 1995). Thus, stimulus-specific persistent neural activity is thought to be a neural substrate for holding memories over short time delays (Major and Tank, 2004). Once persistent activity is triggered within a neuron or neural circuit, its activity can be maintained after the stimulus has terminated. Three general (non-mutually exclusive) mechanisms of persistent activity have been hypothesized: recurrent network activity (Compte et al., 2000; Wang, 2001), short-term synaptic plasticity (Mongillo et al., 2008) and intrinsic biophysical cellular properties. Several studies have demonstrated the role of intrinsic biophysical cellular properties in persistent activity (Egorov et al., 2002; Egorov et al., 2006; Fransen et al., 2006). This firing behavior is linked to cholinergic muscarinic receptor activation and phospholipase C (PLC) signaling in the absence of synaptic reverberations. Two fundamental questions are: (1) What mechanism underlies the generation of sustained firing at a single cell level? (2) What role does intrinsic persistent firing play in working memory? Pharmacological studies suggest that persistent activity relies on activity of Ca2+-activated non-selective cation (CAN) current (ICAN) (Egorov et al., 2002; Egorov et al., 2006). However, the molecules that constitute CAN channels in the brain are not well studied, and the importance of CAN channels to working memory is unknown. I seek to identify molecular mechanisms to convert subthreshold input into intrinsic persistent neural firing in PFC layer 5 pyramidal neurons. I hypothesize that CAN channels are responsible for the intrinsic properties that mediate persistent neural activity in PFC layer 5 neurons. During muscarinic receptor activation, bursts of action potentials will lead to Ca2+ influx. CAN channels will be activated due to the increased intercellular Ca2+ and promote a slow afterdepolarization (sADP), a transition state between subthreshold input and suprathreshold sustained firing. If the sADP is strong enough, it will trigger subsequent spikes, causing further opening of voltage-dependent Ca2+ channels and Ca2+ influx, and thus further opening of CAN channels. Therefore, ICAN will be maintained by a positive feedback loop, generating persistent activity. I have combined electrophysiology, pharmacology, genetics and behavioral analyses to address the potential roles of CAN channels and persistent activity in working memory. First, I confirmed that in the presence of the muscarinic agonist carbachol a brief burst of action potentials triggers a prominent sADP and persistent activity in these neurons. Second, I confirmed that this sADP and persistent firing require activation of a PLC signaling cascade and intracellular calcium signaling. Third, I obtained direct evidence that the transient receptor potential melastatin 5 channel (TRPM5), which is thought to function as a CAN channel in non-neural cells, makes an important contribution to sADP and persistent activity in the layer 5 neurons. Importantly, Trpm5-/- mice show deficits in a Delayed-Non-Match-to-Sample maze (DNMTS) task, a working memory task in the mouse model. Furthermore, PFC-specific expression of TRPM5 using a virally-mediated delivery system in Trpm5-/- mice produced a partial rescue of deficits in the working memory tasks, indicating the importance of TRPM5 in mPFC for performance of these tasks. Lastly, I found that PFC-specific expression of TRPM5 partially rescued the electrophysiological defects in Trpm5-/- mice. By identifying an ion channel contributing to working memory, this work opens the possibility of discovering new drugs for treating working memory deficit.

Memory is a limited resource. Therefore, the circuitry that encodes memory must filter incoming information in accordance with its perceived value. The hippocampus, the hub of the declarative memory system, may achieve memory valuation using its rich variety of neuromodulatory afferent systems. The dopamine (DA) neurons in the ventral tegmental area (VTA) and susbtantia nigra pars compacta (SNpC) are in a particularly strategic position to aid the hippocampus in gating long-term memory. Their firing rates encode the salience of external cues in the environment and they send axons to the output node of the hippocampus, area CA1. In CA1, exogenous receptor stimulation with DA receptor agonists and antagonists suggests an important role for VTA/SNpC DA in learning and memory as the DA receptors powerfully modulate synaptic transmission, permit LTP induction, and enhance different forms of spatial memory. However, it remains unknown whether the VTA/SNpC DAergic axons are capable of activating those receptors and triggering the effects on hippocampal physiology. The VTA/SNpC innervation density in the hippocampus is modest and, in many cases, the axons are distant from the neurons exhibiting the effects. Other sources of DA could couple to those receptors, such as the locus coeruleus, which also releases DA in the CA1 area. To investigate the VTA/SNpC's DAergic influence, I took a circuit-based approach and selectively evoked DA release from the VTA/SNpC DAergic afferents in CA1 in vitro with different patterns of optogenetically guided stimulation. I found that DA release directly modulates the CA3 Schaffer collateral (SC) synaptic excitation of CA1 in a bidirectional manner. A single light-burst (three 5-ms-long pulses at 66 Hz) suppresses the SC-evoked PSP in CA1 pyramidal neurons (PNs) through a D2-receptor dependent enhancement of parvalbumin-positive interneuron mediated feedforward inhibition. More prolonged DA release using 25 light-bursts (at 1 Hz) increases the SC PSP through a D1-type receptor dependent direct presynaptic effect on excitatory transmission. Thus, I propose the following model for how VTA/SNpC DAergic afferents effect oppositional synaptic states to influence learning in the hippocampus in accordance with motivational demands. During tonic DA release, the D4 receptors become activated, globally weaken the SC synaptic input to CA1 PNs, and increase plasticity thresholds. In contrast, phasic DA release activates D1-type receptors, and transitions the SC synapse to a more efficacious state, during which weaker inputs can drive potentiation.

The precision of mammalian movement relies on excitatory sensory feedback supplied by proprioceptors, and its context-dependent refinement by spinal inhibitory microcircuits. One microcircuit that has been implicated in the regulation of sensory input establishes inhibitory synapses directly on the central terminals of sensory neurons. To date, however, the difficulty in gaining selective access to discrete classes of inhibitory interneurons within local microcircuits has left unresolved the contribution of presynaptic inhibition, if any, to motor behavior. Here we have used mouse genetics to gain access to the set of GABAergic interneurons that provide direct input to sensory terminals, and show that their activation evokes the defining physiological features of presynaptic inhibition. Genetic ablation of this set of interneurons in the adult severely perturbs goal-directed reaching movements, and uncovers a pronounced forelimb motor oscillation that appears to have its basis in an enhancement in the gain of sensory feedback. Together, our findings uncover an essential motor behavioral role for this specialized set of presynaptic inhibitory interneurons, and emphasize the relevance of sensory gain control in the neural programming of skilled movement.

Presently, there are no antipsychotic drugs capable of treating the cognitive dysfunctions of schizophrenia. In order to inform the development of better therapies, it is essential to understand the mechanism behind dysfunctional cognition, which requires an understanding of functional cognition. Spatial working memory, a measure of cognitive function, can be assessed in the mouse using a task of delayed alternation: the T-maze. In this thesis, I focus on spatial working memory behavior in the mouse and three brain regions that are implicated in this behavior: the medial prefrontal cortex (mPFC), the hippocampus (HPC) and the medial dorsal thalamus (MD). Lesion and electrophysiological studies in each structure have demonstrated their importance during working memory behavior. Disconnection studies also show that the coordination between the mPFC and either the HPC or MD is important for the behavior, but little is known about the mechanism by which they coordinate. The MD and the ventral region of the hippocampus (vHPC) have robust projections into the mPFC. They are therefore in a good position to influence mPFC activity. Previous reports show that the mPFC and the dorsal region of the hippocampus (dHPC) synchronize activity in the theta range (4-12 Hz) with working memory demand. However, the dHPC does not directly connect with the mPFC so it is unclear how this coordination occurs. We hypothesized that the vHPC may also be involved in spatial working memory behavior and that it may mediate the dHPC-mPFC theta synchrony observed. To test these hypotheses, we recorded neural activity simultaneously from the mPFC, dHPC and vHPC in mice performing the T-maze task. Local field potential oscillations (LFPs), thought to be a measure of synchronized synaptic activity, were obtained from each area. We observed an increase in theta synchrony between the mPFC and both the dHPC and vHPC. Removing the influence of vHPC both analytically and experimentally, we found a decrease in synchrony of the dHPC-mPFC.Aside from the disconnection studies, little is known about the MD-mPFC pathway in rodents. However, due to evidence from schizophrenia patients of altered correlation specifically between the MD and PFC, we hypothesized that an electrophysiological correlate of working memory exists in the MD-mPFC pathway as well and that a decrease in MD activity may lead to prefrontal dysfunction. To test these hypotheses, we recorded LFPs from the mPFC and both single unit activity and LFPs from the MD in mice performing the T-maze task. We observed an increase in phase locking of MD cells to mPFC LFPs in beta (13-30Hz) range during the choice phase of the task. We then utilized a pharmacogenetic technique to decrease firing rate in a small portion of MD cells, which resulted in a deficit in both task acquisition and performance. The increase in MD-mPFC beta phase locking we had observed was not present in MD-inactivated animals. Interestingly, beta coherence between the two structures across learning was highly correlated with choice accuracy on the task. This suggests that MD-PFC coordination is predictive of working memory performance.These findings illustrate how long-range synchrony of the mPFC with HPC in the theta frequency range and with the MD in the beta frequency range may be important markers for normal working memory behavior and if disrupted in humans, could contribute to the cognitive symptoms of schizophrenia.

A stimulus predicting reinforcement can trigger emotional responses, such as arousal, as well as cognitive ones, such as increasing attention towards that stimulus. Neuroscientists have long appreciated that the amygdala mediates spatially non-specific emotional responses, but it remains unclear whether the amygdala links motivational and spatial representations in a way that may be important for the emotional-guidance of attention. To test whether amygdala neurons encode spatial and motivational information, we presented reward-predictive cues in different spatial configurations while assessing whether these cues influenced spatial attention. Cue configuration and predicted reward magnitude modulated amygdala neural activity in a coordinated fashion. Moreover, fluctuations in activity were correlated with trial-to-trial variability in spatial attention. Thus the amygdala integrates spatial and motivational information, which may influence the spatial allocation of cognitive resources. When surveying the environment, animals must be acutely aware of associations between stimuli and aversive outcomes in addition to those resulting in appetitive outcomes. This involves attending to appetitive stimuli in order to obtain positive outcomes, and aversive stimuli in order to avoid negative outcomes. While we first demonstrated that amygdala might play a role in influencing spatial attention towards appetitive stimuli, it is unclear whether the activity of individual amygdala neurons are modulated in a similar way by aversive stimuli that also attract attention. Recording from amygdala neurons while monkeys allocated attention both towards appetitive and aversive stimuli revealed that firing rates reflected where attention was allocated irrespective of valence. We also found that amygdala neurons preferentially encode appetitive and aversive stimuli relative to those of little motivational significance in a conditioning paradigm where spatial characteristics were irrelevant. Thus, amygdala neurons respond with respect to the motivational significance of stimuli, which is tied to spatial attention in contexts involving multiple stimuli. While the amygdala might be involved in guiding attention towards motivationally significant stimuli, this process is likely dependent on its interactions with anatomically linked brain areas. The basal forebrain is a candidate brain area for interacting with the amygdala in influencing emotionally-guided attention given its anatomical connectivity and influence over attentional processes. Here, we analyzed data from amygdala and basal forebrain neurons recorded while spatial attention was captured by appetitive and aversive stimuli. Neurons in the basal forebrain were spatial selective for appetitive and aversive stimuli much like the amygdala. We also found that the timing of value signals differed across brain areas in a manner dependent on the spatial configuration of stimuli. Together, these results demonstrate how the amygdala and basal forebrain may participate in coordinating cognitive and emotional processes and are suggestive of how dysfunction within this pathway might contribute to disorders where emotionally-guided attention is impaired.

Planning behavioral actions requires the ability to form associations between stimuli and outcomes in order to appropriately attribute value and emotional significance to the stimuli. This ability to form associations between stimuli and outcomes is also dependent on being able to attend to the stimulus in question, which generally involves honing in on its spatial location. The amygdala is a brain area that has been investigated extensively in the context of forming associations between stimuli and outcomes; however, whether the amygdala may also be important in linking spatial representations of stimuli with their value is relatively unexplored. Recent work has demonstrated that individual primate amygdala neurons reflect both the value of stimulus-outcome associations and the degree to which spatial attention is directed towards valuable stimuli. While these experiments demonstrated that amygdala neurons are selective for value and spatial information in an attentionally-demanding environment, it is still unclear whether similarly coordinated spatial and value selectivity is present in less attentionally-demanding contexts. To this end, we trained monkeys to perform trace-conditioning tasks similar to those known to induce robust value selectivity within the amygdala; our tasks differed in that we systematically manipulated the spatial location of stimuli in order to evaluate the degree of spatial selectivity in this relatively passive context. Additionally, we used two variants of the trace-conditioning task: a space-irrelevant task in which the relationship between stimuli and outcomes was not dependent on where the stimuli appeared, and a space-relevant task in which the outcome predicted by stimuli was dependent on their spatial location. We reasoned that spatial selectivity in the amygdala might be augmented when spatial variables were relevant to the task, particularly for guiding conditioned responses. This prediction was unsupported, however; amygdala neurons responded similarly in the space-irrelevant and space-relevant tasks. In each task, spatial selectivity was observed mainly around the time that that stimulus was present, and this spatial selectivity was essentially random with respect to neurons' value selectivity. These results run counter to those observed in attentionally-demanding operant tasks, where spatial selectivity was sustained and coordinated with value selectivity, therefore suggesting that spatial coding in the amygdala is task-dependent. Given the weak and unpredictable spatial selectivity in these trace-conditioning tasks, we asked: Under what degree of attentional load are robust spatial signals apparent in the amygdala? To investigate this, we trained monkeys on an operant task where a single stimulus appeared at one of two locations; monkeys had to detect a second stimulus that appeared at the same location, but at an unpredictable time. Unlike in the trace-conditioning tasks, amygdala neurons exhibited sustained spatial selectivity that was well-coordinated with value selectivity on this task. Further suggesting an influential role on attention, the response of amygdala neurons predicted trial-to-trial fluctuations in monkeys' spatial attention. Together, these results show that the amygdala participates in more than just encoding of value-related or emotional stimuli, expanding its role to include encoding of spatial features and lending support to the notion that this brain area may be involved in emotional guidance of spatial attention in physiological and pathological states.

This thesis work is divided into two parts, both guided by the overwhelming evidence that hereditary factors influence neurodevelopment. The first section is focused on advanced paternal age, which may modulate an offspring's place on the continuum of normal behavior, as well as conferring increased risk for the development of certain disorders, such as schizophrenia. By using a mouse model to examine the difference between old and young father offspring, we have been able to flesh out this phenotype. Additionally, by examining the female and male offspring separately, we discovered gender-specific differences between the groups. Ongoing work is seeking to identify changes in methylation between the old and young father offspring that may explain these differences. The second section deals with a specific gene that has been linked to schizophrenia, namely Neuregulin1. This gene plays several roles in neurodevelopement, notably including the proliferation of interneurons and their incorporation into the cortex and olfactory bulb. We used heterozygous mice to explore the effects of a change in gene expression of the proliferation of new neurons from the subventricular zone, their migration through the rostral migratory stream, and differentiation into various interneuron subtypes in the olfactory bulb. The heterozygotes appeared to have decreased turnover of a subset of calretinin-expressing interneurons of the granule cell layer. We also treated subgroups of these mice with clozapine, however this did not seem to have any effect. We looked at the olfactory system in this work since this is a model of neurogenesis that continues into adulthood. Yet the regions that produce cortical interneurons during early development give rise to the subventricular zone. Therefore the findings related to subventricular zone neurogenesis may have similar implications for cortical development.