Effects of Elasticity on Cell Proliferation in a Tissue-Engineering Scaffold Pore.

Scaffolds engineered for in vitro tissue engineering consist of multiple pores where cells can migrate along with nutrient-rich culture medium. The presence of the nutrient medium throughout the scaffold pores promotes cell proliferation, and this process depends on several factors such as scaffold geometry, nutrient medium flow rate, shear stress, cell-scaffold focal adhesions and elastic properties of the scaffold material. While numerous studies have addressed the first four factors, the mathematical approach described herein focuses on cell proliferation rate in elastic scaffolds, under constant flux of nutrients. As cells proliferate, the scaffold pores radius shrinks and thus, in order to sustain the nutrient flux, the inlet applied pressure on the upstream side of the scaffold pore must be increased. This results in expansion of the elastic scaffold pore, which in turn further increases the rate of cell proliferation. Considering the elasticity of the scaffold, the pore deformation allows further cellular growth beyond that of inelastic conditions. In this paper, our objectives are as follows: (i) Develop a mathematical model for describing fluid dynamics, scaffold elasticity and cell proliferation for scaffolds consist of identical nearly cylindrical pores; (ii) Solve the models and then simulate cellular proliferation within an elastic pore. The simulation can emulate real life tissue growth in a scaffold and offer a solution which reduces the numerical burdens. Lastly, our results demonstrated are in qualitative agreement with experimental observations reported in the literature.

Abstinence following intermittent methylphenidate exposure dose-dependently modifies brain glucose metabolism in the rat brain.

Methylphenidate (MP) is a psychostimulant chronically prescribed for the treatment of attention deficit hyperactivity disorder (ADHD). Additionally, MP users may take breaks from using the medication during "drug holidays," which may include short-term or long-term breaks from medication. The present study utilized fluorodeoxyglucose (FDG) positron emission tomography (PET) to analyze the effects of chronic oral MP use and abstinence on brain glucose metabolism (BGluM) in rats at two different doses: high dose (HD) and low dose (LD). The schedule of treatment was 3 weeks on-treatment and 1 week off-treatment for a period of 13 weeks, followed by an abstinence period of 4 total weeks. Results showed that chronic MP treatment using this schedule did not lead to significant changes in BGluM when comparing the control to HD MP groups. However, significant activation in BGluM was observed after periods of abstinence between control and HD MP rats in the following brain regions: the trigeminal nucleus, reticular nucleus, inferior olive, lemniscus, mesencephalic reticular formation, inferior colliculus, and several areas of the cerebellum. These brain regions and functional brain circuit play a role in facial sensory function, the auditory pathway, organizing connections between the thalamus and cortex, motor learning, auditory function, control over eye movement, auditory information integration, and both motor and cognitive functions. These results, when considered with previous studies, indicate that MP schedule of use may have differing effects on BGluM. BGluM following long-term MP use was dependent on MP dose and schedule of use in rats. This study was conducted in non-ADHD model rats with the aim to establish an understanding of the effects of MP itself, especially given the growing chronic off-label and prescribed use of MP. Further studies are needed for analysis of the drug's effects on an ADHD model.

The influence of substrate size upon pulling and gripping forces in parrots (Psittaciformes: Agapornis roseicollis).

The ability to securely grasp substrates of variable diameter is critical to arboreal animals. Arboreal specialists have emerged across several vertebrate lineages - including mammals, lizards and amphibians - and several attempts have been made to quantify their grasping performance, by measuring either gripping (i.e. forces generated about an object or substrate enclosed within the digits) or pulling (i.e. the ability to resist being removed from a substrate) forces. In this study, we present data on both pulling and gripping performance across a range of substrate diameters (0.5-17.5 mm) within a model parrot species (Agapornis roseicollis). Parrots represent an ancient arboreal lineage, allowing us to compare their abilities with those of arboreal specialists within other tetrapod groups. Data were collected using 3D-printed perches of variable diameter, and forces were registered using either an AMTI low-load force plate (grip force) or a Harvard Apparatus portable strength tester (pull force). Gripping forces peaked at a 5 mm diameter perch, while pulling forces were greatest at a 2.5 mm diameter. All forces strongly diminished above 10 mm size, suggesting grip force is optimized when utilizing small perches, a finding which corresponds to observational studies of preferential perching habits among free-ranging parrots. Relative grasping performance (adjusted for body size) in parrots is roughly equivalent to that of other arboreal specialists from other tetrapod lineages, but low when compared with that of raptorial birds that utilize their feet during aerial prey capture. Further taxonomic sampling is encouraged to contextualize how grasping performance varies in an adaptive evolutionary context.

Brief and extended abstinence from chronic oral methylphenidate treatment produces reversible behavioral and physiological effects.

Methylphenidate (MP) is a commonly prescribed psychostimulant to individuals with Attention Deficit Hyperactivity Disorder, and is often used illicitly among healthy individuals with intermittent breaks to coincide with breaks from school. This study examined how intermittent abstinence periods impact the physiological and behavioral effects of chronic oral MP self-administration in rats, and whether these effects persist following prolonged abstinence from the drug. Rats were treated orally with water, low-dose (LD), or high-dose (HD) MP, beginning at PND 28. This daily access continued for three consecutive weeks followed by a 1-week abstinence; after three repeats of this cycle, there was a 5-week abstinence period. Throughout the study, we examined body weight, food intake, locomotor activity, and anxiety- and depressive-like behaviors. During the treatment phase, HD MP decreased body weight, food intake, and depressive- and anxiety-like behaviors, while it increased locomotor activity. During intermittent abstinence, the effects of MP on locomotor activity were eliminated. During prolonged abstinence, most of the effects of HD MP were ameliorated to control levels, with the exception of weight loss and anxiolytic effects. These findings suggest that intermittent exposure to chronic MP causes physiological and behavioral effects that are mostly reversible following prolonged abstinence.

Chronic oral methylphenidate treatment increases microglial activation in rats.

Methylphenidate (MP) is a widely prescribed psychostimulant used to treat attention deficit hyperactivity disorder. Previously, we established a drinking paradigm to deliver MP to rats at doses that result in pharmacokinetic profiles similar to treated patients. In the present study, adolescent male rats were assigned to one of three groups: control (water), low-dose MP (LD; 4/10 mg/kg), and high dose MP (HD; 30/60 mg/kg). Following 3 months of treatment, half of the rats in each group were euthanized, and the remaining rats received only water throughout a 1-month-long abstinence phase. In vitro autoradiography using [3H] PK 11195 was performed to measure microglial activation. HD MP rats showed increased [3H] PK 11195 binding compared to control rats in several cerebral cortical areas: primary somatosensory cortex including jaw (68.6%), upper lip (80.1%), barrel field (88.9%), and trunk (78%) regions, forelimb sensorimotor area (87.3%), secondary somatosensory cortex (72.5%), motor cortices 1 (73.2%) and 2 (69.3%), insular cortex (59.9%); as well as subcortical regions including the thalamus (62.9%), globus pallidus (79.4%) and substantia nigra (22.7%). Additionally, HD MP rats showed greater binding compared to LD MP rats in the hippocampus (60.6%), thalamus (59.6%), substantia nigra (38.5%), and motor 2 cortex (55.3%). Following abstinence, HD MP rats showed no significant differences compared to water controls; however, LD MP rats showed increased binding in pre-limbic cortex (78.1%) and ventromedial caudate putamen (113.8%). These findings indicate that chronic MP results in widespread microglial activation immediately after treatment and following the cessation of treatment in some brain regions.

Mustn1: A Developmentally Regulated Pan-Musculoskeletal Cell Marker and Regulatory Gene.

The Mustn1 gene encodes a small nuclear protein (~9.6 kDa) that does not belong to any known family. Its genomic organization consists of three exons interspersed by two introns and it is highly homologous across vertebrate species. Promoter analyses revealed that its expression is regulated by the AP family of transcription factors, especially c-Fos, Fra-2 and JunD. Mustn1 is predominantly expressed in the major tissues of the musculoskeletal system: bone, cartilage, skeletal muscle and tendon. Its expression has been associated with normal embryonic development, postnatal growth, exercise, and regeneration of bone and skeletal muscle. Moreover, its expression has also been detected in various musculoskeletal pathologies, including arthritis, Duchenne muscular dystrophy, other skeletal muscle myopathies, clubfoot and diabetes associated muscle pathology. In vitro and in vivo functional perturbation revealed that Mustn1 is a key regulatory molecule in myogenic and chondrogenic lineages. This comprehensive review summarizes our current knowledge of Mustn1 and proposes that it is a new developmentally regulated pan-musculoskeletal marker as well as a key regulatory protein for cell differentiation and tissue growth.

Chronic oral methylphenidate treatment reversibly increases striatal dopamine transporter and dopamine type 1 receptor binding in rats.

Previously, we created an 8-h limited-access dual bottle drinking paradigm to deliver methylphenidate (MP) to rats at two dosages that result in a pharmacokinetic profile similar to patients treated for attention deficit hyperactivity disorder. Chronic treatment resulted in altered behavior, with some effects persisting beyond treatment. In the current study, adolescent male Sprague-Dawley rats were split into three groups at four weeks of age: control (water), low-dose MP (LD), and high-dose MP (HD). Briefly, 4 mg/kg (low dose; LD) or 30 mg/kg (high dose; HD) MP was consumed during the first hour, and 10 mg/kg (LD) or 60 mg/kg (HD) MP during hours two through eight. Following three months of treatment, half of the rats in each group (n = 8-9/group) were euthanized, and remaining rats went through a 1-month abstinence period, then euthanized. In vitro receptor autoradiography was performed to quantify binding levels of dopamine transporter (DAT), dopamine type 1 (D1R)-like receptors, and dopamine type 2 (D2R)-like receptors using [3H] WIN35,428, [3H] SCH23390, and [3H] Spiperone, respectively. Immediately following treatment, HD MP-treated rats had increased DAT and D1R-like binding in several subregions of the basal ganglia, particularly more caudal portions of the caudate putamen, which correlated with some previously reported behavioral changes. There were no differences between treatment groups in any measure following abstinence. These findings suggest that chronic treatment with a clinically relevant high dose of MP results in reversible changes in dopamine neurochemistry, which may underlie some effects on behavior.

Identification and Characterization of a Synthetic Osteogenic Peptide.

Osteoporosis is the most common metabolic bone disorder and its management represents a tremendous public health encumbrance. While several classes of therapeutics have been approved to treat this disease, all are associated with significant adverse effects. An algorithm was developed and utilized to discover potential bioactive peptides, which led to the identification of an osteogenic peptide that mapped to the C-terminal region of the calcitonin receptor and has been named calcitonin receptor fragment peptide (CRFP). In vitro treatment of human mesenchymal stem cells with CRFP resulted in dose-specific effects on both proliferation and osteoblastic differentiation. Similarly, in vitro treatment of rat RCJ3.1C5.18 cells led to dose- and species-specific effects on proliferation. A rat ovariectomy (OVX) model was used to assess the potential efficacy of CRFP in treating osteoporosis. MicroCT analysis of distal femoral samples showed that OVX rats treated with CRFP were significantly protected from losses of 55 % in trabecular bone volume fraction (BVF), 42 % in connectivity density, and 18 % in trabecular thickness in comparison to vehicle-treated controls. MicroCT analyses of vertebrae revealed CRFP to significantly prevent a 25 % reduction in BVF. MicroCT evaluation of femoral and vertebral cortical bone found a significant reduction of 2 % in vertebral bone mineral density. In summary, our in vitro studies indicate that CRFP is both bioactive and osteogenic and our in vivo studies indicate that CRFP is skeletally bioactive. These promising data indicate that further in vitro and in vivo evaluation of CRFP as a new treatment for osteoporosis is warranted.

Human nonunion tissues display differential gene expression in comparison to physiological fracture callus.

The healing of bone fractures can become aberrant and lead to nonunions which in turn have a negative impact on patient health. Understanding why a bone fails to normally heal will enable us to make a positive impact in a patient's life. While we have a wealth of molecular data on rodent models of fracture repair, it is not the same with humans. As such, there is still a lack of information regarding the molecular differences between normal physiological repair and nonunions. This study was designed to address this gap in our molecular knowledge of the human repair process by comparing differentially expressed genes (DEGs) between physiological fracture callus and two different nonunion types, hypertrophic (HNU) and oligotrophic (ONU). RNA sequencing data revealed over ∼18,000 genes in each sample. Using the physiological callus as the control and the nonunion samples as the experimental groups, bioinformatic analyses identified 67 and 81 statistically significant DEGs for HNU and ONU, respectively. Out of the 67 DEGs for the HNU, 34 and 33 were up and down-regulated, respectively. Similarly, out of the 81 DEGs for the ONU, 48 and 33 were up and down-regulated, respectively. Additionally, we also identified common genes between the two nonunion samples; 8 (10.8 %) upregulated and 12 (22.2 %) downregulated. We further identified many biological processes, with several statistically significant ones. Some of these were related to muscle and were common between the two nonunion samples. This study represents the first comprehensive attempt to understand the global molecular events occurring in human nonunion biology. With further research, we can perhaps decipher new molecular pathways involved in aberrant healing of human bone fractures that can be therapeutically targeted.

Activation of Wnt signaling in human fracture callus and nonunion tissues.

The Wnt signaling pathway is a key molecular process during fracture repair. Although much of what we now know about the role of this pathway in bone is derived from in vitro and animal studies, the same cannot be said about humans. As such, we hypothesized that Wnt signaling will also be a key process in humans during physiological fracture healing as well as in the development of a nonunion (hypertrophic and oligotrophic). We further hypothesized that the expression of Wnt-signaling pathway genes/proteins would exhibit a differential expression pattern between physiological fracture callus and the pathological nonunion tissues. We tested these two hypotheses by examining the mRNA levels of key Wnt-signaling related genes: ligands (WNT4, WNT10a), receptors (FZD4, LRP5, LRP6), inhibitors (DKK1, SOST) and modulators (CTNNB1 and PORCN). RNA sequencing from calluses as well as from the two nonunion tissue types, revealed that all of these genes were expressed at about the same level in these three tissue types. Further, spatial expression experiments identified the cells responsible of producing these proteins. Robust expression was detected in osteoblasts for the majority of these genes except SOST which displayed low expression, but in contrast, was mostly detected in osteocytes. Many of these genes were also expressed by callus chondrocytes as well. Taken together, these results confirm that Wnt signaling is indeed active during both human physiological fracture healing as well as in pathological nonunions.

The intertwining of transposable elements and non-coding RNAs.

Growing evidence shows a close association of transposable elements (TE) with non-coding RNAs (ncRNA), and a significant number of small ncRNAs originate from TEs. Further, ncRNAs linked with TE sequences participate in a wide-range of regulatory functions. Alu elements in particular are critical players in gene regulation and molecular pathways. Alu sequences embedded in both long non-coding RNAs (lncRNA) and mRNAs form the basis of targeted mRNA decay via short imperfect base-pairing. Imperfect pairing is prominent in most ncRNA/target RNA interactions and found throughout all biological kingdoms. The piRNA-Piwi complex is multifunctional, but plays a major role in protection against invasion by transposons. This is an RNA-based genetic immune system similar to the one found in prokaryotes, the CRISPR system. Thousands of long intergenic non-coding RNAs (lincRNAs) are associated with endogenous retrovirus LTR transposable elements in human cells. These TEs can provide regulatory signals for lincRNA genes. A surprisingly large number of long circular ncRNAs have been discovered in human fibroblasts. These serve as "sponges" for miRNAs. Alu sequences, encoded in introns that flank exons are proposed to participate in RNA circularization via Alu/Alu base-pairing. Diseases are increasingly found to have a TE/ncRNA etiology. A single point mutation in a SINE/Alu sequence in a human long non-coding RNA leads to brainstem atrophy and death. On the other hand, genomic clusters of repeat sequences as well as lncRNAs function in epigenetic regulation. Some clusters are unstable, which can lead to formation of diseases such as facioscapulohumeral muscular dystrophy. The future may hold more surprises regarding diseases associated with ncRNAs andTEs.

Identification of the miRNAome in human fracture callus and nonunion tissues.

Background: Nonunions remain a challenging post-traumatic complication that often leads to a financial and health burden that affects the patient's quality of life. Despite a wealth of knowledge about fracture repair, especially gene and more recently miRNA expression, much remains unknown about the molecular differences between normal physiological repair (callus tissue) and a nonunion. To probe this lack of knowledge, we embarked on a study that sought to identify and compare the human miRNAome of normal bone to that present in a normal fracture callus and those from two different classic nonunion types, hypertrophic and oligotrophic. Methods: Normal bone and callus tissue samples were harvested during revision surgery from patients with physiological fracture repair and nonunions (hypertrophic and oligotrophic) and analyzed using histology. Also, miRNAs were isolated and screened using microarrays followed by bioinformatic analyses, including, differential expression, pathways and biological processes, as well as elucidation of target genes. Results: Out of 30,424 mature miRNAs (from 203 organisms) screened via microarrays, 635 (∼2.1%) miRNAs were found to be upregulated and 855 (∼2.8%) downregulated in the fracture callus and nonunion tissues as compared to intact bone. As our tissue samples were derived from humans, we focused on the human miRNAs and out of the 4223 human miRNAs, 86 miRNAs (∼2.0%) were upregulated and 51 (∼1.2%) were downregulated. Although there were similarities between the three experimental samples, we also found specific miRNAs that were unique to individual samples. We further identified the predicted target genes from these differentially expressed miRNAs as well as the relevant biological processes, including specific signaling pathways that are activated in all three experimental samples. Conclusion: Collectively, this is the first comprehensive study reporting on the miRNAome of intact bone as compared to fracture callus and nonunion tissues. Further, we identify specific miRNAs involved in normal physiological fracture repair as well as those of nonunions. The translational potential of this article: The data generated from this study further increase our molecular understanding of the roles of miRNAs during normal and aberrant fracture repair and this knowledge can be used in the future in the development of miRNA-based therapeutics for skeletal regeneration.

Collagen-Coated Hyperelastic Bone Promotes Osteoblast Adhesion and Proliferation.

Successfully reconstructing bone and restoring its dynamic function represents a significant challenge for medicine. Critical size defects (CSDs), resulting from trauma, tumor removal, or degenerative conditions, do not naturally heal and often require complex bone grafting. However, these grafts carry risks, such as tissue rejection, infections, and surgical site damage, necessitating the development of alternative treatments. Three-dimensional and four-dimensional printed synthetic biomaterials represent a viable alternative, as they carry low production costs and are highly reproducible. Hyperelastic bone (HB), a biocompatible synthetic polymer consisting of 90% hydroxyapatite and 10% poly(lactic-co-glycolic acid, PLGA), was examined for its potential to support cell adhesion, migration, and proliferation. Specifically, we seeded collagen-coated HB with MG-63 human osteosarcoma cells. Our analysis revealed robust cell adhesion and proliferation over 7 days in vitro, with cells forming uniform monolayers on the external surface of the scaffold. However, no cells were present on the core of the fibers. The cells expressed bone differentiation markers on days 3 and 5. By day 7, the scaffold began to degrade, developing microscopic fissures and fragmentation. In summary, collagen-coated HB scaffolds support cell adhesion and proliferation but exhibit reduced structural support after 7 days in culture. Nevertheless, the intricate 3D architecture holds promise for cellular migration, vascularization, and early osteogenesis.

The Use of Hydrogels for the Treatment of Bone Osteosarcoma via Localized Drug-Delivery and Tissue Regeneration: A Narrative Review.

Osteosarcoma is a malignant tumor of bone that leads to poor mortality and morbidity. Management of this cancer through conventional methods involves invasive treatment options that place patients at an increased risk of adverse events. The use of hydrogels to target osteosarcoma has shown promising results both in vitro and in vivo to eradicate tumor cells while promoting bone regeneration. The loading of hydrogels with chemotherapeutic drugs provides a route for site-specific targeted therapy for osteosarcoma. Current studies demonstrate tumor regression in vivo and lysis of tumor cells in vitro when exposed to doped hydrogel scaffolds. Additionally, novel stimuli-responsive hydrogels are able to react with the tissue microenvironment to facilitate the controlled release of anti-tumor drugs and with biomechanical properties that can be modulated. This narrative review of the current literature discusses both in vitro and in vivo studies of different hydrogels, including stimuli-responsive, designed to treat bone osteosarcoma. Future applications to address patient treatment for this bone cancer are also discussed.

Psychostimulants prescribed to children for ADHD following distal radius fractures significantly reduce bone density as a function of duration.

Methylphenidate and mixed amphetamine salts (MAS) are psychostimulant medications widely prescribed for various psychiatric disorders. Although these medications are known to adversely impact bone mineral content and density, as well as biomechanical integrity during skeletal development in rats, their effect on bone density in children remains largely unknown. The primary aim of this work was to investigate the effects of methylphenidate and MAS on bone density following distal radius fractures in pediatric populations, and secondarily assess any impact on healing. The retrospective case-control study was designed to assess fracture healing in patients treated with stimulant drugs and matched controls. For the primary outcome, X-rays (n = 188) were evaluated using an optical density image analysis technique to compare bone density throughout the bone healing process. Results showed that methylphenidate and MAS significantly reduced bone healing by approximately 20% following distal radius fractures in these children. The data also suggested that duration of psychostimulant use played a role in bone healing; the longer the treatment (1-5 years), the lower the bone density was observed (by approximately 52%) as compared to controls (no medication). However, subjects taking these drugs for longer than 5 years did not show a significant difference. Our results suggested that children taking psychostimulants for up to 5 years had slower bone healing following distal radius fractures. Orthopedic surgeons planning elective surgeries should be cognizant of this as a potential issue in recovery after any elective bone procedures and preoperatively optimize bone health as well as counsel patients and their families.

Combined methylphenidate and fluoxetine treatment in adolescent rats significantly impairs weight gain with minimal effects on skeletal development.

Methylphenidate (MP) is frequently prescribed to treat Attention-Deficit/Hyperactivity Disorder (ADHD); however, many patients with ADHD experience depression and anxiety. As such, concomitant administration of selective serotonin reuptake inhibitors such as fluoxetine (FLX) is common. Our laboratory and others have shown that MP impairs skeletal development in preclinical and clinical settings, and FLX has also been linked to skeletal deficits. Unfortunately, little is known about the effects of combined MP and FLX treatment on skeletal development. The objective of this study was to investigate the effects of MP and FLX on bone morphology and biomechanical properties in adolescent rats. Four-week-old male Sprague-Dawley rats were randomly divided into the following 4 groups: Water, MP, FLX, and MP + FLX. As body weights in the MP, FLX, and MP + FLX groups were all lower than Water, the data were compared directly and after adjusting to body weight via linear regression. The direct comparison revealed that MP + FLX rats had significantly shorter (~12 %) and narrower femora and tibiae (~10 %) compared to most other groups, along with shorter (26-35 %), disorganized tibial growth plates. MicroCT analyses of the trabecular compartment of the proximal tibia identified reductions of 47 % for TV, 86 % for BV, 74 % for BV/TV, 68 % for Tb.N, 25 % in Tb.Th, and 74 % in vBMD concomitant with increases of 44 % for Tb.Sp for MP + FLX compared to Water. Similar analyses of femoral midshaft cortical bone identified reductions of 29 % for Ct.V, 30 % for Ps.V, 30 % for Ec. V, and 51 % for pMOI, as well as increases of 17 % for Ct.Th and 2 % for TMD for MP + FLX compared to Water. Biomechanically, MP + FLX femora were weaker, as indicated by a reduction in ultimate force (14 %) in MP + FLX compared to Water. The microstructural and biomechanical effects of MP + FLX were eliminated after adjustment for body weight, though the detrimental effects on growth plate morphology remained. We conclude that while the adverse microstructural and biomechanical effects of MP + FLX seen via direct comparison are predominantly attributable to reductions in body weight rather than direct effects on bone, MP and FLX, particularly in combination show detrimental effects on growth plate structure and chondrocyte morphology. These findings warrant further research into the effect of these drugs on weight gain, skeletal development and growth plate morphology, as well as consideration by physicians treating children and adolescents with ADHD.

Combined Chronic Oral Methylphenidate and Fluoxetine Treatment During Adolescence: Effects on Behavior.

BACKGROUND: Attention Deficit Hyperactivity Disorder (ADHD) can be comorbid with depression, often leading to the prescription of both methylphenidate (MP) and selective serotonin reuptake inhibitor (SSRI) antidepressants, such as fluoxetine (FLX). Moreover, these drugs are often misused as cognitive enhancers. This study examined the effects of chronic oral co-administration of MP and FLX on depressive- and anxiety-like behaviors. METHODS: Adolescent rats received daily either water (control), MP, FLX, or the combination of MP plus FLX in their drinking water over the course of 4 weeks. RESULTS: Data analysis shows a decrease in food consumption and body weight for rats exposed to FLX or the combination of MP and FLX. Sucrose consumption was significantly greater in FLX or MP+FLX groups compared to controls. FLX-treated rats showed no effect in the elevated plus maze (EPM; open arm time) and forced swim test (FST; latency to immobility). However, rats treated with the combination (MP+FLX) showed significant anxiolytic-like and anti-depressive-like behaviors (as measured by EPM and FST), as well as significant increases in overall activity (distance traveled in open field test). Finally, the combined MP+FLX treatment induced a decrease in anxiety and depressive- like behaviors significantly greater than the response from either of these drugs alone. CONCLUSION: These behavioral results characterize the long-term effects of these drugs (orally administered) that are widely co-administered and co-misused and provide important insight into the potential neurobiological and neurochemical effects. Future research will determine the potential risks of the long-term use of MP and FLX together.

Behavioral, Neurochemical and Developmental Effects of Chronic Oral Methylphenidate: A Review.

The majority of animal studies on methylphenidate (MP) use intraperitoneal (IP) injections, subcutaneous (SC) injections, or the oral gavage route of administration. While all these methods allow for delivery of MP, it is the oral route that is clinically relevant. IP injections commonly deliver an immediate and maximum dose of MP due to their quick absorption. This quick-localized effect can give timely results but will only display a small window of the psychostimulant's effects on the animal model. On the opposite side of the spectrum, a SC injection does not accurately represent the pathophysiology of an oral exposure because the metabolic rate of the drug would be much slower. The oral-gavage method, while providing an oral route, possesses some adverse effects such as potential animal injury and can be stressful to the animal compared to voluntary drinking. It is thus important to allow the animal to have free consumption of MP, and drinking it to more accurately mirror human treatment. The use of a two-bottle drinking method allows for this. Rodents typically have a faster metabolism than humans, which means this needs to be considered when administering MP orally while reaching target pharmacokinetic levels in plasma. With this oral two-bottle approach, the pathophysiological effects of MP on development, behavior, neurochemistry and brain function can be studied. The present review summarizes these effects of oral MP which have important implications in medicine.

Chronic oral methylphenidate plus fluoxetine treatment in adolescent rats increases cocaine self-administration.

Background: Depression and attention deficit hyperactivity disorder are known to be comorbid. Treatment of these commonly coexisting diseases typically involves the combined prescription of methylphenidate (MP), a psychostimulant, and fluoxetine (FLX), a selective serotonin reuptake inhibitor (SSRI). MP and cocaine have similar mechanisms of action and this study examined the effects of chronic treatment of MP combined with FLX on cocaine consumption in rats. Methods: Four groups of rats received access to drinking solutions of water (control), MP (30/60 mg/kg/day), FLX (20 mg/kg/day), or the combination of MP (30/60 mg/kg/day) plus FLX (20 mg/kg/day), during 8 h per day for one month. Following these drug treatments, rats were allowed to self-administer cocaine for 14 days. Results: Our results showed that, during the first week of cocaine self-administration, the MP-treated rats had significantly greater numbers of active lever presses (plus 127%) and increased consumption of cocaine compared to the control rats. In contrast, during week two of cocaine self-administration, the rats treated with the MP + FLX combination showed significantly more lever presses (plus 198%) and significantly greater cocaine consumption (plus 84%) compared to the water controls. Conclusion: Chronic oral treatment during adolescence with the combination of MP plus FLX resulted in increased cocaine use after 2 weeks of cocaine self-administration in rats. These novel findings suggest that the combined exposure to these two drugs chronically, during adolescence, may produce increased vulnerability towards cocaine abuse during young adulthood.