Browsing by Author "Baijnath, Sooraj."

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Evaluation of the pharmacodynamic effects of Ketamine on neurotransmitter levels and CREB/BDNF expression in rat brain.
(2020) Khoza, Leon Joseph.; Baijnath, Sooraj.
Mental disorders contribute to 13% of the global burden of disease. With major depressive disorder (MDD) expected to be the most significant contributor by 2030, the economic and social impact of this burden will be substantial. There have been various factors linked to the underlying pathophysiology of MDD, including a deficit in individual vital neurotransmitter connections between specific neurons, and alterations in the expression of the transcription factors cyclic AMP response element-binding protein (CREB) and the brain-derived neurotrophic factor (BDNF) in the brain. Ketamine, an N-methyl-D-aspartate receptor (NMDAR) rantagonist and an 􀄮-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) stimulator has officialy been used as an anesthetic and analgesic drug since the 1970s, until recently when it was approved for the management and treatment of MDD. Ketamine works by altering and rebalancing the monoaminergic, glutamatergic and GABArgic sytems to stimulate new synaptic connections, better memory, and improved brain plasticity. However, there are limited published studies that demonstrate the direct relationship between ketamine, brain neurotransmitters levels, gene and protein expression in the management of MDD. In this study, we investigated the pharmacodynamic effects of ketamine in the brain by assessing changes in monoaminergic, glutaminergic and GABAergic neurotransmitter levels using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Furthermore, we investigated the links between ketamine and the expression of transcription factors, cyclic AMP response element-binding protein (CREB) and brain-derived neurotrophic factor (BDNF) in treatment of depression using quantitative real-time polymerase chain reaction (qPCR) analysis. Twenty-one healthy male Sprague-Dawley (SD) rats were administered 15􀯗mg/kg of ketamine via intraperitoneal administration at different time points (N = 3 per time point). Experimental animals were euthanized by decapitation post-administration of ketamine, and brain samples were harvested for analysis. As per LC-MS/MS and qPCR, the pharmacodynamic results demonstrated that ketamine􀂶s anti-depressive mechanism of action is due to alteration of the glutamatergic system which occurs via the disinhibition of glutamate release, which further boosts central nervous synaptogenesis, hence maintaining the in-balance neurotransmitters and genes associated with the pathophysiology of depression.
Evaluation of the pharmacokinetics of ketamine for the treatment of major depressive disorder.
(2018) Naidoo, Vivian Campbell.; Baijnath, Sooraj.; Naicker, Tricia.; Naidoo, Panjasaram.; Kruger, Hendrik Gerhardus.
Recent reports have demonstrated ketamine’s potential use in the treatment of major depressive disorder (MDD), as it elicits potent antidepressant effects via a different mechanism compared to conventional antidepressants. Ketamine’s hypothesized antidepressant effect is elicited by a neurochemical cascade involving the antagonization of the N-methyl-D-aspartate (NMDA) receptors and the subsequent activation of the α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors; resulting in the disinhibition of glutamate signalling due to the suppression of tonic glutamate input into the GABAergic interneurons, providing rapid symptomatic relief as opposed to the two-week delay with conventional treatments. There is a large escalation in the number of individuals being diagnosed with treatment resistant depression (TRD) even after numerous trials on conventional antidepressant therapy. Health care professionals are now resorting to unconventional treatments, such as ketamine’s off-label use, to achieve therapeutic outcomes and provide symptomatic relief. MDD’s increasing prevalence has been associated with significant public health costs and morbidity rates and therefore alternative, effective treatments are now essential. Many reports have been published on the intranasal (IN) efficacy of ketamine in the treatment of major depressive disorder, however there have been no studies investigating the effects on the route of administration in drug delivery to the brain. The purpose of this study was to investigate pharmacokinetics of ketamine following oral, intraperitoneal and intranasal administration. A dose of 15mg/kg (body weight) was administered to healthy male Sprague-Dawley rats, and ketamine concentrations were quantified in both plasma and brain tissue homogenates at time intervals of 5, 15, 30, 60, 120, 240 minutes post-treatment. The results showed that with intraperitoneal administration, concentrations of 524,58 ng/mL and 352,06 ng/mL, were achieved in plasma and brain tissue, respectively. Surprisingly, IN administration which is believed to favour drug delivery to the brain only exhibited moderate levels post administration; whereas, oral administration produced significantly lower levels due to extensive first-pass metabolism of ketamine in the liver and intestines. These results show that parenteral administration should be used for the administration of ketamine in the treatment of MDD. The findings of the study provide a platform for future investigations assessing alternative routes of administration of ketamine; and its use in clinical practice for the treatment of MDD. This paves the way forward to optimize treatment and provide symptomatic relief were conventional antidepressants have failed those suffering with MDD.
Evaluation of the use of cannabidiol in the treatment of anxiety related disorders by assessing changes in neurotransmitter levels and expression of CREB/BDNF in the rodent brain.
(2020) Haripershad, Advaitaa Meera.; Baijnath, Sooraj.; Naicker, Tricia.
Anxiety and anxiety-related disorders are common psychiatric disorders that are responsible for high disease burden. The pathogenesis of anxiety involves dysfunction in the limbic brain regions including the amygdala, prefrontal cortex, and hippocampus. The current pharmacological treatments for anxiety target the modulation of the activity monoamine neurotransmitters such as dopamine, serotonin, gamma-aminobutyric acid, norepinephrine and glutamate. These neurotransmitters are key in the regulation of the maladaptive responses of anxiety. Primary pharmacotherapies demonstrate limitations in drug efficacy as well as adverse side effects, highlighting the need for novel therapeutics for anxiety and anxiety-related disorders. Cannabidiol (CBD), a non-psychoactive cannabinoid from the Cannabis sativa plant, has been considered a potential anxiolytic treatment as a result of its interaction in the endocannabinoid system, which regulates synaptic plasticity and neuronal activity implicated in the anxiety response. The therapeutic potential of CBD against neuropsychiatric disorders have been reported in preclinical and clinical studies. Since the global increase in cannabis legalization, there remains a need to supplement the available literature related to the neural effect of cannabis use on behavioural, neurochemical and biochemical changes. There are gaps in the knowledge of the pharmacokinetics and behavioural effects of CBD. This study will contribute to increasing the knowledge of the effect of cannabis on neurotransmitters and molecular changes in the brain. In this thesis, chapter 1 is a literature review focusing on the neurobiology and pharmacological treatments of anxiety, cannabidiol as a treatment for anxiety, and the neurotransmitters and genes implicated in anxiety. In addition to this, chapter 1 also reviews the theory of the experimental processes performed in this study. Chapter 2 is the publication “Evaluation of the use of cannabidiol in the treatment of anxiety related disorders by assessing changes in neurotransmitter levels and expression of CREB/BDNF in the rodent brain” submitted to The Journal of Neuroscience Research. Chapter 3 is the summary and conclusion of the thesis.
Functional and molecular changes associated with intranasal buprenorphine in a healthy rodent model.
(2020) Xhakaza, Sanelisiwe Penelope.; Baijnath, Sooraj.
Opioid addiction is a spiralling global epidemic associated with intense drug craving and the compulsive use of opiate drugs such as heroin, oxycodone, oxymorphone amongst others. Buprenorphine (BUP), commercially available as Subutex, is a partial opioid agonist that is used to treat opioid addiction and pain. It is associated with minimal risks of overdose and can be used outside of clinical care, making it the safest and most preferred choice of drug in the treatment of opioid addiction, over methadone and naltrexone. Literature suggests that opioids carry out their effects by altering the neurotransmitter systems of the brain viz. dopamine, norepinephrine, serotonin, glutamate and gamma-aminobutyric acid. Therefore, an ideal treatment drug should be able to counter these neurotransmitter changes in the brain. There is currently a lack of information on the pharmacodynamic effects of BUP in the brain, more specifically on how the drug affects brain neurotransmitter levels and its effect on the transcription factors Brain-derived neurotrophic factor (BDNF) and Cyclic AMP Response Element-Binding Protein (CREB). This study evaluates the pharmacokinetics of BUP, its effect on neurotransmitter levels and the expression of BDNF and CREB at various time points following a single dose. Sprague-Dawley rats received 36 μL of 0.3 mg/mL of BUP via intranasal administration. Following dosing, animals were euthanised and brain tissues were collected at different time points. A rapid and sensitive liquid chromatograph-mass spectrometry (LC-MS method was developed for the quantification of BUP and neurotransmitters (dopamine, serotonin, glutamate, norepinephrine and gamma-aminobutyric acid) in brain tissue and the expression of CREB and BDNF was determined using qPCR. This thesis is divided into three chapters. Chapter 1 contains a thorough background on BUP, opioid addiction and the role of neurotransmitters, BDNF and CREB. It also explains the principles of the quantification techniques used in this study i.e LC-MS and qPCR. Chapter 2 is a manuscript that was submitted to Addiction Biology titled “Functional and molecular changes associated with intranasal buprenorphine administration in a healthy rodent model”. Lastly, Chapter 3 provides a general conclusion and future recommendations for the study. The results in this present study indicate that BUP leads to significant changes in neurotransmitters, CREB and BDNF over time. Providing a better understanding of the mechanism of action of the drug, which could possibly improve the treatment of opioid addiction.
In vitro and in vivo evaluation of metal-chelating agents as novel metallo beta-lactamase inhibitors against carbapenem-resistant enterobacteriaceae.
(2018) Omolabi, Kehinde Foluke.; Baijnath, Sooraj.; Kruger, Hendrik Gerhardus.
Infectious diseases remain one of the leading causes of death worldwide, despite the discovery of new and improvements on existing antibiotics. Bacteria are constantly developing sophisticated mechanisms of resisting the effects of antibiotics, this in turn has increased their pathogenicity and virulence. Drugs belonging to the beta-lactam class of antibiotics are most commonly prescribed as they display a broad-spectrum activity against both gram-positive and gramnegative bacteria. Carbapenems which are a member of this class is regarded as the last line of defence against bacterial infections. Resistance to carbapenems is on the increase especially by bacterial strains that are capable of producing metallo-beta lactamase enzymes. Infections caused by carbapenem resistant Enterobacteriaceae are deadly especially those mediated by metallo beta-lactamases. Efforts are being made to synthesize compounds that can inhibit these enzymes. Thus far little progress has been made as a clinically available metallo beta-lactamase inhibitor has not yet emerged, hence the scourge of carbapenem resistant infections rages on. Therefore, the main aim of this study was to evaluate the in vitro and in vivo activities of metal chelating agents NO3PY and NOTA as potential metallo beta-lactamase inhibitors against carbapenem resistant Enterobacteriaceae. The metal-chelating agents used in this study were NOTA and NO3PY. In vitro analysis was performed to determine the minimum inhibitory concentrations by broth microdilution of meropenem alone and when co-administered with the chelators against resistant bacterial strains. The strains used in this study were Escherichia coli NDM-1, Klebsiella pneumoniae 449, Escherichia coli IMP-1 and Enterobacter cloacae NDM-1. Time kill kinetics was also evaluated at graded concentrations of MIC, 1*MIC, 2*MIC, 4*MIC, 8*MIC and 16*MIC. For the in vivo pharmacokinetics were determined using LC-MS/MS analysis. Forty-eight healthy male Balb/c mice were divided into two groups; meropenem+NO3PY group and meropenem+NOTA group. Both groups received intraperitoneal doses at 10 mg/kg of meropenem and the MBLIs. Thereafter, the in vivo efficacy of meropenem co-administered with NOTA (100 mg/kg each) in a murine thigh infection was determined. Both chelators were able to restore the efficacy of meropenem to a concentration as low as 0.06 µg/ml. The time kill kinetics also showed that both compounds were able to significantly extend the killing time of meropenem. In vivo pharmacokinetic analysis revealed that NO3PY may not xiv be a suitable candidate for in vivo efficacy study as the MBLI was not bioavailable in plasma at 10mg/kg. NOTA on the other hand was bioavailable at the same concentration as NO3PY. The former was able to potentiate the effect of meropenem in vivo in a murine thigh infection model. It was evident by a significant reduction of colony forming unit counts in groups treated with meropenem co-administered with NOTA when compared to infected controls Further preclinical work such as in vitro and in vivo cytotoxicity tests, post beta-lactamase inhibitor effects among others are recommended for NOTA to further ascertain its suitability as a potential clinical metallo beta-lactamase inhibitor.
LC-MS/MS method development and validation for simultaneous quantification of first-line HIV drugs and second-line TB drugs in rat plasma.
(2018) Malinga, Thembeka Hlengiwe.; Govender, Thavendran.; Baijnath, Sooraj.; Naicker, Tricia.; Kruger, Hendrik Gerhardus.
Tuberculosis (TB) and human immunodeficiency virus (HIV) co-infection continues to be a major public health concern, worldwide. HIV infection has increased the TB incidence over the past twenty years, making it hard to eliminate TB. At the same time, TB continues to be responsible for approximately 30% of deaths among HIV-infected individuals. Emtricitabine (FTC), efavirenz (EFV), and tenofovir (TFV) are constituents of a one-day-pill, AtriplaTM, which was approved in 2006 by the Food and Drug Administration (FDA). AtriplaTM is a triple combination anti-HIV drug that provides an efficient dosing plan. Streptomycin (STR), kanamycin (KAN), and ofloxacin (OFL) are second-line anti-TB drugs used to treat multidrug-resistant/ extensively drug-resistant tuberculosis (MDR/XDR-TB). The worldwide increase in the prevalence of anti-TB drugs resistance is of concern to researchers since it remains one of the most significant threats to the community. Co-prescription of anti-HIV and anti-TB drugs poses a challenge of drug-drug interactions, which causes adverse effects. Therapeutic drug monitoring (TDM) seems to be the tool for a solution to these problems since it personalizes doses thus reducing drug toxicity. LC-MS/MS methods with short run times are required to produce effective TDM studies. Therefore, this study aimed to evaluate the new Ascentis Express column technologies [pentafluorophenylpropyl (F5), octadecyl (C18), biphenyl, and reversed phase amide (RP-Amide)] and their applicability to the simultaneous quantification of current first-line anti-HIV treatment Atripla. It also aimed to develop, optimize and validate a liquid-chromatography tandem mass spectrometry (LC-MS/MS) methods for the simultaneous quantification of anti-HIV drugs (FTC, EFV, and TFV) and second-line anti-TB drugs (STR, KAN, and OFL) in rat plasma for the usage of TDM. The currently used HPLC columns have longer run times making them impractical in a point of care environment since the number of patients and diseases is globally increasing. There are also no or very few studies regarding the LC-MS/MS method of simultaneous HIV and TB drugs for HIV positive TB patients. The biphenyl column showed consistency and optimum performance with regard to the number of theoretical plates, resolution and peak asymmetry factor. It showed good separation and overall effectiveness. However, this does not rule out other columns for other purposes intended to be accomplished. The LC-MS/MS method developed for the simultaneous quantification of anti-HIV drugs and second-line anti-TB drugs was short to eleven minutes and met all the recommendations by the European Medicines Agency (EMA) guidelines for bioanalytical method validation. The new HPLC column matrices (F5, C18, biphenyl, and RP-Amide) offer various benefits such as the potential of saving solvents and short runtimes, essential in TDM studies. Therefore, the usage of the new HPLC column technologies iv will be beneficial in a point of care environment in terms of saving time and money. The LC-MS/MS method validated in this study can be used in clinical trials and in the simultaneous determination of the effective plasma concentrations of anti-TB and anti-HIV drugs, making it a strong candidate for TDM in a point of care setting.
Mass spectrometric imaging for Tuberculosis drug development.
(2017) Baijnath, Sooraj.; Govender, Thavendran.; Naicker, Tricia.; Kruger, Hendrik Gerhardus.
For many years, Tuberculosis (TB) has plagued the human race claiming millions, if not billions, of lives. With the advent of short-course chemotherapy TB has become a manageable disease, however in recent times Mycobacterium tuberculosis has developed resistance to a number of established and trusted antibiotics. This coupled with severe forms of extra-pulmonary TB, has placed significant emphasis on the development of new anti-TB agents. The drug development process is a long and costly affair, with less than 1% of new drugs reaching clinical trials. This is where molecular imaging, in particular mass spectrometry imaging (MSI), is fast becoming a promising tool in the evaluation of drug candidates. MSI can be used to streamline the drug development process by fast tracking areas of target identification, target quantification, pharmacokinetics, drug distribution and tissue localization. MSI possesses some distinct advantages in terms of sample preparation and the lack of the need for radiolabeling, making it the ideal technique for in vivo tissue drug distribution studies. The objectives of this study were to demonstrate the value of MSI in the development and evaluation of new and existing TB antibiotics, focusing on central nervous system (CNS) manifestations of the disease. In order to achieve these objectives, two of the most promising antimycobacterial agents, clofazimine (CFZ) and linezolid (LIN), were selected. Initially, the distribution of these agents in a healthy animal model was investigated, since these would represent the minimum tissue concentrations achievable. The single-dose study for both drugs were similar, in that there was poor penetration into the brain after a 100mg/kg dose in a healthy murine and rodent model, respectively. A four-week multiple dose study was conducted, each of the antibacterials showed excellent accumulation in the CNS, with preference to specific areas of the brain, demonstrating the neuroprotective potential of these drugs (Chapters 2 and 3). For the effective evaluation of anti-TB drugs, the lung has to be taken into consideration since this is the primary site of M.tb infections. However, the lung poses problems in terms of sample preparation for MSI. Since the lung is responsible for gaseous exchange, it is made up of a number of air-filled spaces that are kept “open” by a fine balance in pressure, inside and outside the lung. When this balance is disturbed, such as when the thoracic cavity is pierced, to collect tissue, the lung collapses. This results in distortion of tissue structure and subsequent distribution information can be misleading. For this reason, we evaluated various established cryoprotectants as lung inflation media. This inflation procedure would main structural integrity of the lung and provide accurate tissue distribution data. From the cryoprotective agents tested in this experiment we found that 10% DMSO was ideal, in terms of structural preservation and accurate drug distribution (Chapter 4). As part of this series of experiments other anti-bacterial agents were also evaluated, to demonstrate the value of MSI in drug development. These drugs also appear in the antibiotic pipeline; tetracyclines, tigecycline (TIG) and doxycycline (DOX), rifampicin (RIF), gatifloxacin (GAT) and pretomanid (PA-824). The findings were very interesting in that each agent displayed a unique pattern of distribution, this is due to the chemical nature of these drugs and their interaction with the blood-brain-barrier (BBB). In addition to this, we have demonstrated how MSI can be used to determine various aspects of drug-tissue interaction for drug development. MSI was used to prove that the chemical properties of a drug do not always govern its movement across the BBB. RIF is a large drug molecule that one would not expect to permeate the brain, however this experiment has demonstrated its time-dependent distribution in the brain (Chapter 5). The results show how the tetracyclines have widespread tissue distribution in the brain, which contributes to their efficacy in the treatment of brain damage (Chapters 6 and 7). This technique was also used to understand how GAT enters the brain and contributes to the proven neurotoxicity of the flouroquinolones (Chapter 8). In the final chapter, we showed how MSI can be used in the tissue evaluation of novel antibiotics, such as pretomanid (Chapter 9). These findings emphasize the need to evaluate the drug distribution of antibiotics, since pathogens manifest themselves in different areas of the brain and cause damage. This information will be invaluable in our pursuit of effective treatments to CNS diseases and disorders, allowing medical practitioners to develop more targeted treatment programmes.