Traumatic injuries to the spinal cord (SCI) and peripheral nerve (PNI) affect several thousand people worldwide every year. At present, there is no effective treatment for SCI and despite continuous improvements in microsurgical reconstructive techniques for PNI, many patients are still left with permanent, devastating neurological dysfunction. This thesis investigates the effects of mesenchymal stem cells (MSC) derived from adipose (ASC) and dental (DSC) tissue and chitosan/microRNA-124 polyplex particles on regeneration after spinal cord and peripheral nerve injury in adult rats. Dental stem cells were obtained from apical papilla, dental pulp, and periodontal ligament. ASC and DSC expressed MSC surface markers (CD73, CD90, CD105 and CD146) and various neurotrophic molecules including BDNF, GDNF, NGF, VEGF-A and angiopoietin-1. Growth factor stimulation of the stem cells resulted in increased secretion of these proteins. Both ASC and DSC supported in vitro neurite outgrowth and in contrast to Schwann cells, ASC did not induce activation of astrocytes. Stimulated ASC also showed an enhanced ability to induce capillary-like tube formation in an in vitro angiogenesis assay. In a peripheral nerve injury model, ASC and DSC were seeded into a fibrin conduit, which was used to bridge a 10 mm rat sciatic nerve gap. After 2 weeks, both ASC and DSC promoted axonal regeneration in the conduit and reduced caspase-3 expression in the dorsal root ganglion (DRG). ASC also enhanced GAP-43 and ATF-3 expression in the spinal cord, reduced c-jun expression in the DRG and increased the vascularity of the implant. After transplantation into injured C3-C4 cervical spinal cord, ASC continued to express neurotrophic factors and laminin and stimulated extensive ingrowth of 5HT-positive raphaespinal axons into the trauma zone. In addition, ASC induced sprouting of raphaespinal terminals in C2 contralateral ventral horn and C6 ventral horn on both sides. Transplanted cells also changed the structure and the density of the astroglial scar. Although the transplanted cells had no effect on the density of capillaries around the lesion site, the reactivity of OX42-positive microglial cells was markedly reduced. However, ASC did not enhance recovery of forelimb function. In order to reduce activation of microglia/macrophages and the secondary tissue damage after SCI, the role of microRNA-124 was investigated. In vitro transfection of chitosan/microRNA-124 polyplex particles into rat microglia resulted in the reduction of reactive oxygen species and TNF-α levels and lowered expression of MHC-II. Upon microinjection into uninjured rat spinal cords, particles formed with Cy3-labeled control sequence RNA, were specifically internalized by OX42 positive macrophages and microglia. Alternatively, particles injected in the peritoneum were transported by macrophages to the site of spinal cord injury. Microinjections of chitosan/microRNA-124 particles significantly reduced the number of ED-1 positive macrophages after SCI. In summary, these results show that human MSC produce functional neurotrophic and angiogenic factors, creating a more desirable microenvironment for neural regeneration after spinal cord and peripheral nerve injury. The data also suggests that chitosan/microRNA-124 particles could be potential treatment technique to reduce neuroinflammation.

Trophic factors have shown great promise in their potential to treat neurological disease. In particular, glial cell line-derived neurotrophic factor (GDNF) has been identified as a potent neurotrophic factor for midbrain dopamine (DA) neurons in the substantia nigra (SN), which lose function in Parkinson’s disease (PD). GDNF progressed to phase II clinical trials, which did not meet proposed endpoints. The large size and binding characteristics of GDNF have been suspected to contribute to some of the shortcomings of GDNF related to delivery to target brain regions. Smaller peptides derived from GDNF (Dopamine-Neuron Stimulating Peptides – DNSPs) have been recently investigated and appear to demonstrate trophic-like effects comparable to GDNF. In the described studies, a time course study was conducted to determine in vivo DA-release characteristics 1-, 2- and 4- weeks after peptide treatment. These studies determined the effects on DA terminals within striatal sub-regions using microelectrodes. A heterogeneous effect on striatal sub-regions was apparent with the maximum effect in the dorsal striatum – corresponding to terminals originating from the SN. Dysregulation of GDNF or GDNF signaling is believed to contribute to motor dysfunction in aging and PD. Thus, it is hypothesized that GDNF is necessary for the maintenance and function of neurons. To extend this line of investigation, in vivo functional measures (DA-release and -uptake) and behavioral and cellular alterations were investigated in a transgenic mouse model (Gdnf+/-) with reduced GDNF protein levels. The described studies determined that both DA-uptake and -release properties were altered in middle-aged Gdnf+/- mice with only modest reductions in DA neurochemical levels. GDNF levels in Gdnf+/- mice were restored to levels comparable to wild-type (WT) counterparts by treatment with GDNF. GDNF protein supplementation led to enhanced motor behavior and increased markers for DA neurons in the SN of Gdnf+/- mice. Gdnf+/- mice appeared to show a heightened sensitivity to GDNF treatment compared to WT counterparts. Overall, this body of work examines novel synthetic peptides with potential to enhance DA-neuron function and expands upon the current understanding of GDNF’s role in the nigrostriatal pathway.

Major depressive disorder (MDD) and bipolar disorder (BD) are psychiatric diseases that affect behavior and impair cognition. A gene important to these disorders is the brain derived neurotrophic factor (BDNF) which is involved in processes controlling neuroplasticity. Previous studies have suggested that BDNF expression levels have to be finely regulated for normal mental health and cognition. This study therefore aimed to identify cis-regulatory elements that regulate BDNF promoter IV (BP4), which plays a role in mood and cognition, and investigated how polymorphisms in these cis-regulatory elements might alter BP4 activity contributing to MDD and BD. BP4-LacZ transgenic mice and primary neuron cultures were used to show that BP4 was active in the hippocampus, cortex and amygdala and responded to PKC, KCl and Wnt signaling activation. Using comparative genomics, two highly conserved regions were identified, BE5.1 and BE5.2, which contain the rs10767664 and rs12273363 polymorphisms respectively. Reporter gene assays in primary cultures derived from these brain structures showed that BE5.1 and BE5.2 were responsible for “filtering” or “gating” the effects of different combination of activated signal transduction pathways on BP4. Thus, BE5.1 increased BP4 response to forskolin in cortical cultures while abolishing BP4 response to PMA in hippocampal cultures. Similarly, BE5.2 permitted BP4 response to KCl and combined forskolin and PMA treatment, but not individual forskolin and PMA treatment nor LiCl in cortical cultures. Significantly, the minor allele of rs12273363, which has been associated with MDD and BD susceptibility, acted as a more potent repressor of BP4 response to neuron depolarization by KCl and PKA/PKC activation in different primary cultures. The possible relevance of these findings to the role of altered BDNF expression in MDD and BD are discussed.

Stress is a major risk factor for substance abuse. Intermittent social defeat stress increases drug self-administration (SA) and elevates brain-derived neurotrophic factor (BDNF) expression in the ventral tegmental area (VTA) in rats. Intra-VTA BDNF overexpression enhances social defeat stress-induced cross-sensitization to psychostimulants and induces nucleus accumbens (NAc) ΔFosB expression. Therefore, increased VTA BDNF may mimic or augment the development of drug abuse-related behavior following social stress. To test this hypothesis, adeno-associated virus (AAV) was infused into the VTA to overexpress either GFP alone (control) or GFP + BDNF. Rats were then either handled or exposed to intermittent social defeat stress before beginning cocaine SA training. The SA acquisition and maintenance phases were followed by testing on a progressive ratio (PR) schedule of cocaine reinforcement, and then during a 12-h access "binge" cocaine SA session. BDNF and ΔFosB were quantified postmortem in regions of the mesocorticolimbic circuitry using immunohistochemistry. Social defeat stress increased cocaine intake on a PR schedule, regardless of virus treatment. While stress alone increased intake during the 12-h binge session, socially-defeated rats that received VTA BDNF overexpression exhibited even greater cocaine intake compared to the GFP-stressed group. However, VTA BDNF overexpression alone did not alter binge intake. BDNF expression in the VTA was also positively correlated with total cocaine intake during binge session. VTA BDNF overexpression increased ΔFosB expression in the NAc, but not in the dorsal striatum. Here we demonstrate that VTA BDNF overexpression increases long-access cocaine intake, but only under stressful conditions. Therefore, enhanced VTA-BDNF expression may be a facilitator for stress-induced increases in drug abuse-related behavior specifically under conditions that capture compulsive-like drug intake.

Schwermut, Einsamkeit, Desinteresse: 2-7% der Weltbevölkerung werden von diesen Gefühlen bestimmt, sie sind an einer Depression erkrankt. In Deutschland leiden aktuell bis zu 4 Mio. Menschen an einer Depression. Zwei Drittel diese Kollektivs befinden sich in hausärztlicher Behandlung, aber nur etwa die Hälfte von diesen Patienten wird korrekt diagnostiziert und behandelt. Die Gründe hierfür sind auf der einen Seite die schwierige Diagnosefindung, auf der anderen die bisher nicht vollständig geklärte Ätiologie und die komplexen Wirkmechanismen der medikamentösen antidepressiven Therapieoptionen. Die Entstehung einer depressiven Episode ist multifaktoriell bedingt, wobei insbesondere genetischen Faktoren in der Pathogenese depressiver Erkrankungen eine wichtige Rolle zugeordnet wird. Neurobiologische Untersuchungen der molekularen und biochemischen Hintergründe von depressiven Episoden befassen sich zurzeit insbesondere mit drei Hypothesen: Diese sind die Neurotransmitter-Dysbalance-Hypothese (Monoaminhypothese), die Stressachsen-Hypothese (Hypothalamus-Hypophysen-Achse) und die Neurotrophin-Hypothese. Die Stressachsen-Hypothese befasst sich in diesem Zusammenhang mit der Aktivität des Hypothalamus-Hypophysen-Systems, die bei depressiv erkrankten Patienten dysreguliert ist und mit einer erhöhten Kortisolsekretion einhergeht. Die Monoaminhypothese der Depression postuliert eine Dysfunktion serotonerger, noradrenerger und dopaminerger Systeme. Die Neurotrophinhypothese bezieht sich auf das Second Messenger-System des durch Antidepressiva aktivierten cyclischen Adenosin-Monophosphat (cAMP) Signalwegs. Cyclisches Adenosin-Monophosphat aktiviert die cAMP-abhängige Proteinkinase (PKA), die wiederum den Transkriptionsfaktor cAMP response element binding protein (CREB) phosphoryliert und ihn somit stimuliert. Aktiviertes CREB verstärkt die Transkription zahlreicher Zielgene, inklusive die des brain-derived neurotrophic factor (BDNF), welcher unter anderem als Regulator von Neurotransmittern dient und Überleben, Differenzierung und Plastizität von Neuronen beeinflusst. Astrozyten gehören zur Gruppe der Makrogliazellen im zentralen Nervensystem (ZNS) und sind die größte Population der Gliazellen. Sie sind für die Kaliumhomöostase und ebenso für die Regulation der synaptischen Transmission und der neurovaskulären Koppelung zuständig. Des Weiteren scheinen Astrozyten eine wichtige Rolle für die Bildung glialer Narben, die Induktion der Blut-Hirn-Schranke und auch für das neuronale Überleben zu spielen. Bei der Analyse der Wirkmechanismen medikamentöser antidepressiver Therapien ist in der letzten Zeit die Rolle der Astrozyten in den Vordergrund gerückt, um deren Beitrag zu antidepressiven Therapien zu untersuchen. Das Ziel der vorliegenden Arbeit war zu untersuchen, ob in Astrozyten der Serotonin-Transporter (5-HT-Transporter, 5-HTT), der Brain-derived neurotrophic factor (BDNF), der Dopamin-Transporter oder die Stickstoffmonoxyd-Synthase III (endotheliale Stickstoffmonoxid-Synthase, eNOS) gebildet werden und falls ja, ob sich deren Expression durch Applikation von Antidepressiva ändert. Die aus Rattenhirnen gewonnenen Astrozytenkulturen wurden hierfür entweder mit dem trizyklischen Antidepressivum Imipramin, dem selektiven Serotoninrückaufnahmeinhibitor Escitalopram oder zur Kontrolle mit Kochsalzlösung inkubiert. Nachdem die entsprechende mRNA aus den behandelten Astrozytenkulturen extrahiert wurde, ist sie in cDNA translatiert und mit Hilfe der quantitativen Real-Time PCR quantifiziert worden. Durch Vergleich der Expression des zu untersuchenden Gens mit der Expression der Housekeeping-Gene 18s-rRNA, Glycerinaldehyddehydrogenase (GAPDH) und Acidic ribosomal phosphoprotein (ARP) wurden Ungenauigkeiten bei der cDNA-Synthese ausgeglichen und die Daten normalisiert. Die rechnerische Auswertung der quantitativen Real-Time PCR erfolgte unter Verwendung der Ct-Werte unter Zuhilfenahme der geNORM Software. Die Ergebnisse zeigen eine signifikant erhöhte BDNF-Expression nach Imipramingabe. Hierbei zeigen bei den getrennten Untersuchungen der jeweiligen mRNA Chargen die mit 100µM Imipramin behandelten Astrozytenkulturen stärker signifikante Ergebnisse, als die mit 50µM Imipramin behandelten Astrozytenkulturen. Werden alle Proben, die mit der jeweiligen Imipraminkonzentration 4 Stunden inkubiert wurden, zusammen analysiert und mit den jeweiligen Kontrollen verglichen, zeigt sich sowohl bei der Behandlung mit 50µM Imipramin als auch mit 100µM Imipramin eine signifikante Steigerung der BDNF Expression. Escitalopram stimulierte die BDNF-Expression zwar ebenfalls nominell, jedoch war der Effekt nicht signifikant. Des Weiteren konnte eine deutliche Expression von 5-HTT-mRNA in Astrozytenkulturen nachgewiesen werden. Jedoch hatte keines der Antidepressiva einen signifikanten Effekt auf die 5-HTT-Expression. Es konnten weder in den behandelten, noch in den unbehandelten Astrozytenkulturen DAT oder NOS-III nachgewiesen werden. / Melancholia, loneliness, lack of interest: 2-7% of world population are ruled by these feelings. In Germany, 4 million inhabitants suffer from depression. Two third of this population are under medical treatment, but only half of these patients are diagnosed and medicated correctly. Reasons are difficulties in finding the exact diagnosis on the one hand and the vague aetiology and complex effects of antidepressants on the other. The origin of a depressive episode has multiple reasons, especially genetic components are considered to play an important role. Neurobiological investigations of the molecular and biochemical backgrounds of depressive episodes centre on three hypotheses: The dysbalance hypothesis of neurotransmitters (monoamine hypothesis), the hypothesis of activated hypothalamus-pituitary-adrenal axis and the neurotrophin hypothesis. In this context, the hypothesis of activated hypothalamus-pituitary-adrenal axis deals with the dysregulation and over-stimulation of the hypothalamus-pituitary-adrenal axis and the following elevated cortisol-secretion. The monoamine hypothesis of depressive episodes postulates a dysfunction of serotonergic, noradrenergic and dopaminergic systems. The neurotrophin hypothesis refers to the cyclic adenosine monophosphate (cAMP) second messenger system, activated by antidepressants. Cyclic adenosine monophosphate activates the cAMP dependent protein kinase (PKA) that phosphorylates and consequently stimulates the transcription factor cAMP response element binding protein (CREB). Activated CREB enhances the transcription of numerous genes, including the brain derived neurotrophic factor (BDNF) that is among others regulating neurotransmitters and influences neuronal survival, differentiation and plasticity. Astrocytes are part of the macroglia in central nervous system and form the major population of glial cells. They are considered to be responsible for homeostasis of potassium and the regulation of synaptic transmission and neurovascular linkage. Furthermore, astrocytes take part in forming glial cicatrixes, induction of blood-brain-barrier and neuronal survival. In the course of investigating the effects of antidepressants, the centre of interest has focused on astrocytes and their contribution to antidepressant therapy. The aim of this dissertation was to study, if the serotonin transporter (5-HTT), the brain derived neurotrophic factor (BDNF), the dopamin transporter or the nitric oxide synthase III (eNOS) is synthesised in astrocytes and in case of, if their expression is modulated by antidepressants. The astrocyte cultures, gained from rats’ brains, were incubated with the tricyclic antidepressant imipramine, the selective serotonin reuptake-inhibitor escitalopram or sodium chloride solution. After extracting the corresponding mRNA from the astrocyte cultures, it was translated to cDNA and quantified using the quantitative Real-Time PCR. Comparing the expression of the gene of interest with the expression of the housekeeping genes 18s-rRNA, glyceraldehydes phosphate dehydrogenase (GAPDH) and acidic ribosomal phosphoprotein (ARP), equated inexactness of cDNA synthesis and normalised the acquired data. The mathematical evaluation of the qRT-PCR based on the Ct-figures with the assistance of geNORM software. The results show a significantly elevated BDNF expression after imipramine administration. mRNA batches of astrocyte cultures incubated with 100µM imipramine presented more significant results than mRNA batches of astrocyte cultures incubated with 50µM imipramine. Analysing all samples incubated for four hours with the corresponding concentration of imipramine and compared with the sodium chloride solution controls, a significant elevation of BDNF expression is detected in 50µM imipramine samples as well as in 100µM imipramine samples. Escitalopram stimulates the expression of BDNF nominally without showing significant effects. Furthermore, a clear expression of 5-HTT-mRNA in astrocyte cultures could be detected, but none of the antidepressants induced a significant effect on 5-HTT expression. Neither in medicated nor in control samples DAT or NOS-III were detected.

The primary purpose of this study was to investigate the effects of aerobic and resistance training on BDNF and IGF-I expression in a 3xTg-AD mouse model of Alzheimer’s disease. Twenty-four 3xTg-AD mice were randomly assigned to either an aerobic (AT, n=8), resistance (RT, n=8), or control (CNT, n=8) group. Intervention groups underwent 9 weeks of exercise training. Motor behavior and grip strength were measured pre- and post- intervention. Our results showed a significant increase in hippocampal BDNF expression in AT mice after a 9-week intervention. Further, AT mice were found to have higher concentrations of IGF-I, and improved motor behavior when compared to RT and CNT. No significant differences were observed in IGF-I concentration between RT and other groups. RT improved grip strength after nine weeks of training. These findings support the use of AT and RT as a tool to improve comorbidities found in Alzheimer’s disease. / Includes bibliography. / Thesis (M.S.)--Florida Atlantic University, 2018. / FAU Electronic Theses and Dissertations Collection

Ependymin (EPN) is a neurotrophic factor (NTF) that functions in goldfish long-term memory formation and optic nerve elongation (Shashoua, 1976; Shashoua, 1977; Shashoua, 1985). Goldfish EPN, or CMX-8933 (a short goldfish EPN mimetic studied by our lab), surprisingly have several demonstrated effects on mammalian cells, including neuroregenerative effects in a rat stroke model (Shashoua et al, 2003), and the activation of therapeutic superoxide dismutase (SOD) (Parikh, 2003) and transcription factor AP-1 (Adams et al, 2003) in mouse neuroblastoma cells or rat primary cortical neuronal cultures. Among its various functions, AP-1 can function as a master switch in long-term memory consolidation (Sanyal et al, 2002), so it may be a key event in EPN's mechanism of action. AP-1 activation is also a characteristic associated with other full-sized neurotrophic factors, including nerve growth factor and brain-derived nerve growth factor. This thesis was divided into three parts. The purpose of part I was to determine whether our previously observed upregulation of SOD by CMX-8933 is dependent upon (or merely concurrent with) AP-1 activation. Four independent SOD immunoblot experiments demonstrated that pre-treatment of rat primary cortical cultures with trifluoromethyl pyrimidine carboxylate (TFPC), a specific inhibitor of AP-1, significantly (p = 0.0004) decreased cellular levels of SOD by 67% at its IC50 concentration of 1 ìM, and completely inhibited the upregulation at 10 and 100 ìM concentrations. Thus, the CMX-8933-induced upregulation of SOD appears to depend (directly or indirectly) on AP-1 activation. Part II of this thesis included the use of bioinformatics to re-verify exciting recent observations that EPN-like proteins exist in mammals, termed mammalian-ependymin-related proteins or MERPs (Apostolopoulos et al, 2001). If our analyses were convincing, human EPN mimetics would then be designed and tested for AP-1 activation. Computer alignments and hydropathy plots performed with EPN amino acid sequences deduced from gene entries in GenBank verified the existence of mammalian homologs containing highly conserved domains with fish EPN's, suggesting the possibilities of similar protein conformation and function. Two human EPN mimetics were designed, hEPN-1 (8 aa long, corresponding to the same region as CMX-8933) and hEPN-2 (14 aa long, containing CMX-8933 and 6 upstream aa). Several mimetic doses were tested on mouse Neuro-2a cultures for nuclear translocation of c-Jun and c-Fos proteins (comprising the AP-1 particle upregulated by fish CMX-8933). Seven independent c-Jun immunoblot experiments, and five c-Fos experiments, demonstrated a strong (as high as 25-fold) dose-dependent increase in the nuclear titers of the AP-1 proteins. Both peptides had statistically equivalent effects. Thus, human EPN appears to exist, and two mimetics derived from its sequence appear to be biologically active against mouse neuroblastoma cells. Since hEPN-1 and -2 have only a few residues in common with CMX-8933, we hypothesize that the mimetic shape rather than sequence may be important for biological activity. In part III of this thesis, the biological effects of hEPN-1 and hEPN-2 on mouse Neuro-2a cells were studied further using RT-PCR to analyze potential increases in specific mRNAs. mRNAs related to growth, energy production, and protein translation were tested since previous data in our lab (Kaska, 2003) indicated mRNAs for translational elongation factor-2 (EF-2), and ribosomal proteins L19 and S12 were upregulated in rat primary cortical cultures by fish mimetic CMX-8933 (Kaska, 2003). Treatment of Neuro-2a cells with 1.0 ìg/ml hEPN-1 (the highest dose tested for the AP-1 translocation experiments) for 24 hrs appeared to increase (N = 1) mRNAs for ATP Synthase-C, ribosomal protein L19, and translational EF-2, relative to the levels of housekeeper polyubiquitin. Thus hEPN-1 may be involved in processes related to growth. Altogether, the data from this thesis extends our knowledge of fish EPN mimetic CMX-8933 (showing that its induction of SOD requires AP-1), demonstrates that human EPN may exist (bioinformatics), shows that two human EPN mimetics are biologically active (induce AP-1 translocation), and shows that one mimetic hEPN-1 may activate several mRNAs related to growth in mouse Neuro-2a cells.

The purpose was to analyze the role of dopamine D1 and D2 receptors in conditioned place preference (CPP) of a relatively high dose (5 mg/kg) of methylphenidate (MPH) in adolescent male and female rats, as well as the role of these receptors in the effects of MPH on brain-derived neurotrophic factor (BDNF). The primary mechanism of MPH in the brain is the blockade of the dopamine transporter, yielding an increase of dopamine in the synaptic cleft and is the basis for the rewarding properties of MPH. An initial preference given on postnatal day (P)32 yielded no preference for any context in a three-chambered shuttle box with removable dividers, thus, a biased procedure was used. Conditioning began the day after the initial preference test on P33. On conditioning trials, animals were first administered saline or their respective antagonist (D1 antagonist: 0.1 or 0.2 mg/kg SCH-23390; D2 antagonist: 0.01 or 0.03 mg/kg Eticlopride HCl), followed by methylphenidate (MPH; 5mg/kg). Approximately 10 min after MPH administration, rats were placed into the paired context for a 10 min trial. The choice of the paired context was balanced across animals. In a separate session, all animals received saline in the opposing context. One day post-conditioning on P38, a preference test was administered with dividers removed. Preference was determined through the amount of time spent in the paired context as compared to time spent in the unpaired context on the post-conditioning preference test. One day following the preference test on P39, brain tissue was removed, and nucleus accumbens and striatum analyzed for BDNF. Results showed that MPH produced an increased preference on the post-conditioning preference test that was blocked by either dose of SCH-23390, but was not affected by either dose of eticlopride. Additionally, the higher dose of SCH-23390 resulted in a conditioned place aversion in males, which may be due to the increased presence of dopamine D1 receptors in adolescent males. MPH produced a robust significant increase in BDNF in both nucleus accumbens and striatum, and this increase was alleviated by SCH-23390, but the effect on BDNF is still to be analyzed relative to D2 antagonism. These results show that MPH results in a conditioned place preference in adolescent male and female rats, and these effects appear to be mediated by the dopamine D1 receptor. Further, MPH results in a significant increase of BDNF in drug reward areas of the brain, which has implications towards synaptic plasticity in these regions in response to MPH.

This study analyzed the role of dopamine D1 and D2 receptors in methylphenidate (MPH) conditioned place preference (CPP) in adolescent male and female rats, in addition to the role of these receptors in the effects of MPH on brain-derived neurotrophic factor (BDNF) in the dorsal striatum and nucleus accumbens. Using a nonbiased CPP procedure, the animals were conditioned from postnatal day (PD) 33 to 37. On conditioning trials, animals were first administered saline or their respective antagonist (0.1 or 0.2 mg/kg SCH-23390; 0.01 or 0.03 mg/kg eticlopride HCl), followed by MPH (5 mg/kg). Approximately 10 min after MPH administration, the rats were placed into the paired context for a 10-min trial. One day after conditioning on PD38, a preference test was administered with dividers removed. One day following the preference test on PD39, brain tissue was removed, and the nucleus accumbens and striatum were analyzed for BDNF. Results revealed that MPH conditioning resulted in an increased preference that was blocked by either dose of SCH-23390, but generally not affected by either dose of eticlopride. Further, the higher dose of SCH-23390 resulted in a conditioned place aversion in males, presumably due to an increased number of dopamine D1 receptors in adolescent males. MPH produced a significant increase of striatal and accumbal BDNF alleviated by SCH-23390 or eticlopride. These results show that MPH results in CPP in adolescent male and female rats and these effects appear to be mediated by the dopamine D1 receptor, but the effects of MPH on BDNF appear to be mediated by both dopamine receptor families.

Methylphenidate (MPH) is a psychostimulant that is used for the treatment of attention-deficit hyperactivity disorder that is often recreationally abused. Past studies have primarily analyzed the effects of MPH on behavior and BDNF using males as subjects, with studies showing a lack of behavioral sensitization, although the effect of MPH on BDNF has yielded contradictory results. BDNF is a neurotrophin ubiquitously found throughout the brain that plays an important role in synaptic maintenance and development and has been implicated in addiction. This study analyzed sex differences in induction and expression of MPH locomotor sensitization in adolescent male and female rats as well as the effects of MPH on BDNF protein in two brain areas of drug reward: the nucleus accumbens (NAcc) and striatum (STR), after both induction and expression of sensitization. After habituation to a locomotor arena, animals (N=6-8 per group) were administered MPH (5 mg/kg) or saline every other day from postnatal day (P)33 to 49 and tested for 30 min in the same arena with activity counts recorded. In one group, brain tissue was removed one day following testing and the NAcc and STR assayed for BDNF at P50. A different group of animals was raised to P60 and given an MPH (or saline) challenge. One day following the challenge, brain tissue was removed and the NAcc and STR were assayed for BDNF at P61. Females administered MPH demonstrated behavioral sensitization from P33 to P41, and then decreased in activity from P41 to P49. Females demonstrated a robust increase in locomotor activation as compared to males, which failed to demonstrate sensitization to MPH. However, both groups given MPH demonstrated an increase in activity compared to controls throughout sensitization testing. On the challenge at P60, females administered MPH demonstrated higher levels of activity compared to all other groups and were equivalent to their final day of sensitization. Males administered MPH also expressed sensitization, as they demonstrated increased behavioral activation as compared to saline-treated controls. Neurochemical analyses at P50 revealed that MPH produced a significant increase in striatal BDNF in males, but a significant decrease in striatal BDNF in females. There were no changes in the NAcc. At P61, BDNF was increased in both STR and NAcc in males, and female data will be presented. These data demonstrate robust sex differences in behavioral activation and sensitization to MPH that is both induced and expressed in females, but only expressed in males. Further, MPH produces sex-dependent effects on BDNF, indicating sex differences in the brain plasticity response to MPH in adolescence.