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Title:  Identification, diagnosis, and treatment of neuropathologies, neurotoxicities, tumors, and brain and spinal cord injuries using microelectrodes with microvoltammetry
United States Patent: 
7,112,319
Issued: 
September 26, 2006

Inventors: 
Broderick; Patricia A. (Bronx, NY), Pacia; Steven V. (New York, NY)
Assignee: 
The Research Foundation of the City University of New York (New York, NY), New York University (New York, NY)
Appl. No.: 
10/118,571
Filed: 
April 8, 2002


 

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Abstract

The present invention relates to devices and methods of use thereof for making semiderivative voltammetric and chronoamperometric measurements of chemicals, e.g. neurotransmitters, precursors, and metabolites, in vitro, in vivo, or in situ. The invention relates to methods of diagnosing and/or treating a subject as having or being at risk of developing a disease or condition that is associated with abnormal levels of one or more neurotransmitters including, inter alia, epilepsy, diseases of the basal ganglia, athetoid, dystonic diseases, neoplasms, Parkinson's disease, brain injuries, spinal cord injuries, and cancer. The invention provides methods of differentiating white matter from grey matter using microvoltammetry. In some embodiments, regions of the brain to be resected or targeted for pharmaceutical therapy are identified using Broderick probes. The invention further provides methods of measuring the neurotoxicity of a material by comparing Broderick probe microvoltammograms of a neural tissue in the presence and absence of the material.

SUMMARY OF THE INVENTION

The present invention relates to devices and methods for microvoltammetric and/or chronoamperometric imaging of temporal changes in neurotransmitter concentrations in living humans and non-human animals comprising contacting cells with a Broderick probe, applying a potential to said Broderick probe, and generating a temporally resolved microvoltammogram. The method may further comprise determining from said microvoltammogram the presence and concentration of at least one marker selected from the group consisting of serotonin, dopamine, ascorbic acid, norepinephrine, .gamma.-aminobutyric acid, glutamate, neurotensin, somatostatin, dynorphin, homovanillic acid, uric acid, tryptophan, tyrosine, nitrous oxide, and nitric oxide. Methods of the invention may further comprise comparing the microvoltammogram and/or neurotransmitter concentrations to reference a microvoltammogram and/or neurotransmitter concentration(s).

The present invention relates to devices and methods of use thereof for determining the presence and concentration of chemicals in a cell, tissue, organ or organism. The invention relates to, inter alia, semiderivative voltammetric measurements and chronoamperometric measurements of chemicals, e.g. neurotransmitters, precursors, and metabolites. The invention relates to methods of diagnosing and/or treating a subject as having or being at risk of developing a disease or condition that is associated with abnormal levels of one or more neurotransmitters including, inter alia, epilepsy, diseases of the basal ganglia, athetoid, dystonic diseases, neoplasms, Parkinson's disease, brain injuries, spinal cord injuries, and cancer. Microvoltammetry methods of the invention may be performed in vitro, in vivo, or in situ. The invention provides methods of differentiating white matter from grey matter using microvoltammetry. In some embodiments, the invention provides methods of brain tumor diagnosis using distinct white matter voltammetric signals as detected by Broderick probes. The methods of the invention may be applied to cancer diagnosis and treatment, even where a tumor has infiltrated other tissue, e.g. white matter. In some embodiments, the invention provides methods of treating a tumor wherein the tumor's position is determined using microvoltammetry. The invention relates to use of Broderick probes to determine the concentration of a material, e.g. dopamine, norepinephrine, and serotonin, in the brains of patients having epilepsy. In some embodiments of the invention, regions of the brain to be resected are identified using Broderick probes. In some embodiments of the invention, regions of the brain to be targeted for pharmaceutical therapy are identified using Broderick probes. The invention further provides methods of measuring the neurotoxicity of a material by comparing Broderick probe microvoltammograms of a neural tissue in the presence and absence of the material.

The present invention relates to devices and methods for treating epilepsy. More specifically, the invention relates to use of Broderick probes to ascertain neurotransmitter levels in the brains of patients having epilepsy, especially temporal lobe epilepsy. In some embodiments of the invention, regions of the brain to be resected are identified using Broderick probes. In some embodiments of the invention, regions of the brain to be targeted for pharmaceutical therapy are identified using Broderick probes.

The present invention also relates to devices and methods for reliably distinguishing temporal lobe gray matter from white matter using Broderick probes with microvolatammetry. The invention further relates to methods of brain cancer diagnosis using distinct white matter voltammetric signals as detected by Broderick probes. The invention further relates to diagnosis of other white matter diseases. Nonlimiting examples of white matter diseases are multiple sclerosis, leukodystrophies, mitochondrial diseases, lipid disorders and glial cell-related disorders whether these glial cells or glia are normal, abnormal, modified or cultured and the like.

The present invention further relates to devices and methods for diagnosing and treating cocaine psychomotor stimulant behaviors. In some embodiments of the invention microelectrodes may be contacted with a subject to ascertain changes in neurotransmitter levels, e.g. due to release and/or reuptake, in real time. In some embodiments, the invention provides methods of predicting the occurrence of movement disorder effects of a drug. Nonlimiting examples of movement disorders are cocaine addiction, Huntington's disease, Parkinson's disease, Autism, Lesch-Nyhan Disease and the like.

The present invention further provides devices and methods for diagnosing pathologies and/or abnormalities of neurotransmitter levels. Neurotransmitters that may be detected by the techniques of the invention may be selected from the group consisting of serotonin (5-HT), dopamine (DA), ascorbic acid (AA), norepinephrine (NE), .gamma.-aminobutyric acid (GABA), glutamate, neurotensin, somatostatin, dynorphin, homovanillic acid, uric acid (UA), tryptophan, tyrosine, nitrous oxide, and nitric oxide.

DETAILED DESCRIPTION OF THE INVENTION

Broderick probe--a microelectrode comprising graphite, oil, and a material selected from the group consisting of glycolipids, lipoproteins, saturated and unsaturated fatty acids, and perfluorosulfonated materials. A non-limiting example of a Broderick probe may be found in FIG. 1. Further details and examples may be found in U.S. Pat. Nos. 4,883,057, 5,443,710, and 5,938,903, all to P. A. Broderick including circuit diagrams and methods of making Broderick probes. Broderick probes may be in electrical contact with an auxiliary electrode and/or a reference electrode. It will be apparent to those of ordinary skill in the art, particularly in view of the cited patent documents, that "Broderick probe" is a term that relates to a number of microelectrodes that vary by composition and the type of circuit in which it is employed and that these variations give rise to differences in detection properties.

Broderick probes are miniature carbon-based sensors that are able to detect electrochemical signals for a vast number of neurotransmitters, neuromodulators and metabolites, including neuropeptides, hormones and vitamins (Broderick P A, 1989, U.S. Pat. No. 4,883,057; Broderick P A, 1995, U.S. Pat. No. 5,433,710; Broderick P A, 1997, EP 0487647 B1; Broderick P A, 1999, U.S. Pat. No. 5,938,903; Broderick P A, 1999, Hong Kong, HK # 1007350). These probes have made it possible to routinely and selectively detect in discrete neuroanatomic substrates of living human and animal brain, the monoamines, DA, NE, and 5-HT, in addition to the precursor to 5-HT, 1-tryptophan (1-TP), ascorbic acid (AA) and uric acid (UA) (Broderick P A, 1988, Neurosci. Lett. 95:275 280; Broderick P A, 1989, Brain Res. 495:115 121; Broderick P A, 1990, Electroanalysis 2:241 251; Broderick P A, 2000, Epilepsia 41(Suppl.):91; Broderick P A et al., 2000, Brain Res. 878:49 63). It is also possible to differentiate catecholamines, DA and NE, electrochemically using these probes (Broderick P A, 1988, Neurosci. Lett. 95:275 280; Broderick P A, 1989, Brain Res. 495:115 121; Broderick P A, 1990, Electroanalysis 2:241 251; Broderick P A, 2000, Epilepsia 41(Suppl.):91; Broderick P A et al., 2000, Brain Res. 878:49 63). More recently, it has been found that these probes are also capable of electrochemical detection of somatostatin and dynorphin A (Broderick P A, 2000, Epilepsia 41(Suppl.):91).

In some embodiments of the invention, the Broderick probe is a Broderick probe microelectrode as shown in the schematic diagram in FIG. 1. Within the field of electrochemistry, this sensor is termed the indicator microelectrode and is also called the working microelectrode. The surface of the microelectrode consists of carbon-base and is the electrochemical device.

Broderick probes do not promote bacterial growth either before or after sterilization with gamma irradiation. Gamma irradiation treatment was performed by Sterigenics International, Inc., Haw River, N.C.

Broderick probes can be used effectively for different applications in human and animal surgery. Preliminary studies with Broderick probe stearic versus lauric acid microelectrodes in vitro, in situ, and in vivo showed a possible advantage for the lauric acid microelectrodes for use short-term, e.g., intraoperative recordings, and a possible advantage for stearic acid for use long-term, e.g., chronic monitoring in humans and animals (Broderick P A, 1989, U.S. Pat. No. 4,883,057; Broderick P A, 1995, U.S. Pat. No. 5,433,710; Broderick P A, 1997, EP 0487647 B1; Broderick P A, 1999, U.S. Pat. No. 5,938,903; Broderick P A, 1999, Hong Kong, HK #1007350; Hope O et al., 1995, Cocaine has remarkable nucleus accumbens effects on line, with behavior in the serotonin-deficient Fawn Hooded rat. NIH/NIGMS Symposium, Washington, D.C.).

Broderick probes can detect basal (normal, natural, endogenous or steady state) concentrations of neurotransmitters and other neurochemicals in vivo, in situ and in vitro. They can also detect alterations in these neurotransmitters or neurochemicals in brain, or body before and after pharmacological manipulation with drugs or other compounds. Neurochemicals during actual, induced or even mimicked brain diseases can be detected as well. Example 5 focuses on 5-HT alterations in NAcc in freely moving animals during normal open-field behaviors of locomotor (exploratory) and stereotypy compared with, in the same animal, cocaine psychomotor stimulant effects on 5-HT and behavior.

Changing the surface of the sensor changes the capacitance of the surface of the sensor. The surface of the indicator microelectrode is a capacitance diffuse double layer (C.sub.dt) that allows potential to accumulate on its surface. Capacitance is a critical aspect of charging (background) current. Charging current is a current pulse that flows through the C.sub.dt to allow faradaic electron transfer to begin. Accumulation of potential on the surface of the indicator microelectrode is necessary for faradaic electron transfer. Charging current is proportional to electrode surface area; therefore, these miniature sensors (200 microns and less in diameter) minimize charging current effects.

Broderick probes can be used in conjunction with classical electrical circuits used in electrochemistry such as chronoamperometry, differential pulse voltammetry and double differential voltammetry. Another electrical circuit for providing an output signal having a mathematical relationship in operation to an input signal can be semiderivative or semidifferential. These two terms are used interchangeably here, although these two circuits have some technical differences. Semiderivative electroanalysis diminishes non-faradaic current by the addition of analysis time. In the present studies, a CV 37 detector (BAS, West Lafayette, Ind.) was equipped with a semiderivative circuit. This circuit uses a linear scanning methodology as its basis. Semiderivative treatment of voltammetric data means that the signals are recorded mathematically as the first half derivative of the linear analog signal. A semiderivative circuit combines an additional series of resistors and capacitors, called a "ladder network" (Oldham,K, 1973, Anal. Chem. 45:39 50) with the traditional linear scanning technology which then allows more clearly defined waveforms and peak amplitudes of electrochemical signals than was previously possible with linear scanning methodology.

Broderick probe microvoltammogram--These may be plotted as current versus time or as current versus applied potential. Other renderings are also possible. The concentration of biogenic amines and other materials may be deduced from these microvoltammograms, e.g. according to the Cottrell equation. According to the invention, a microvoltammogram is broadly defined as any rendering of the signals from a Broderick probe susceptible to human perception including, but not limited to, paper, electronic, and virtual representations of the Broderick probe signal. An individual of sufficient skill in the art to perceive a Broderick probe signal in real-time, e.g. from a visual display screen, is also within the contemplation and scope of this definition.

The main strength of in vivo microvoltammetry (electrochemistry) is that it allows the study of the neurochemical time course of action of normal neurochemistry, as well as the neurochemistry after an administered drug regimen. Temporal resolution is fast, in seconds and milliseconds. Moreover, the attendant microspatial resolution is superior (availability of discrete areas of brain without disruption). Both highly sensitive temporal and spatial resolution makes these studies ultimately most efficient for mechanism of action studies Another strength lies in the fact that these in vivo microvoltammetric studies are done in the freely moving and behaving animal model, using the same animal as its own control (studies in the living human brain are underway as well). Thus, a direct determination of whether or not a neurochemical effect is abnormal can be made because the normal neurochemical effect is seen a priori.

The basic in vivo electrochemistry experiment involves the implantation of an indicator electrode in a discrete and specified region of brain, the application of a potential to that electrode, the oxidation or reduction of the selected neurochemical and the recording of the resultant current. In essence, the potential is applied between the indicator and the reference electrode; the reference electrode provides a relative zero potential. This is an electrochemical technique with which information about an analyte, a neurotransmitter, or its metabolite, including its concentration, is derived from an electrochemical current as a function of a potential difference. This potential difference is applied to the surface of an electrochemical electrode.

In microvoltammetry, each neurotransmitter, metabolite, precursor to neurotransmitter, etc. is identified by the peak oxidation potential, or half-wave potential at which the neurochemical generates its maximum current. Using the Broderick Probe stearic acid microelectrode inserted in NAcc, the oxidation potential at which DA generates its maximum current in vivo (physiological pH, 37.5.degree. C.) was empirically determined to be +0.140 V (SE .+-.0.015 V) in over one thousand studies. The oxidation potential at which serotonin generates its maximum current under the same conditions was empirically determined to be +0.290 V (SE.+-.0.015 V) in over one thousand studies.

What matters in microvoltammetry is that each of these biogenic amines have amine groups that are protonated at neutral pH and therefore, exist as cations, whereas metabolites of the monoamines are deprotonated at neutral pH and exist as anions (Coury L A et al., 1989, Biotechnology 11:1 37). Thus, the monoamine metabolites such as the metabolites of DA, 3, 4 dihydroxyphenylacetic acid, (DOPAC), 3,4-dihydroxyphenylglycol (DHPG-DOPEG) and homovanillic acid (HVA) cannot interfere with the detection of DA at the same peak oxidation potential or half-wave potential, characteristic for DA.

The same principles are applicable to detection of the biogenic amine, 5-HT. Serotonin is detected without interference at the same oxidation potential or half-wave potential from either its metabolite, 5-hydroxyindoleacetic acid (5-HIAA) or UA, which is a constituent of brain with similar electroactive properties to those of 5-HT. Factors such as the significantly lower sensitivity of the indicator microelectrode to anions, the charge and diffusion characteristics of each catecholamine or indoleamine vis-a-vis its metabolites, preclude such interference. Descriptions of each neurochemical detected by this inventor with Broderick probes are published in detail (Broderick P A, 1995, U.S. Pat. No. 5,433,710; Broderick P A, 1996, EP 90914306.7; Broderick P A, 1999, U.S. Pat. No. 5,938,903; Broderick P A, 1989, Brain Res. 495:115 121; Broderick P A, 1988, Neurosci. Lett. 95:275 280; Broderick P A, 1990, Electroanalysis 2:241 245; Broderick P A, 1993, Pharmacol. Biochem. Behav. 46:973 984; Broderick P A, 2002, Handbook of Neurotoxicology, Vol. 2, Chapter 13; Broderick P A et al., 2000, Brain Res. 878:48 63; Broderick P A et al., 1997, Neuroscience and Biobehavioral Reviews 21(3):227 260; Broderick P A, 1989, U.S. Pat. No. 4,883,057; Broderick P A, 1997, EP 0487647 B1; Broderick P A, 1999, Hong Kong, HK #1007350; Broderick P A, 2000, Epilepsia 41(Suppl.):91).

An important distinction between the detection of signals in microvoltammetry as compared with the detection of signals in microdialysis is that in microvoltammetry, the indicator microelectrode is the detecting device, whereas in microdialysis methods, the dialysis membrane is a membrane and not the detecting device. The microdialysis membrane is simply a membrane through which perfusate is collected. The perfusate is then brought to the high performance liquid chromatography (HPLC) device, equipped with an electrochemical column that is the actual detecting device. These electrochemical columns range in millimeters in diameter, whereas microvoltammetry indicator microelectrodes range from single digit microns to a few hundred microns in diameter.

A common misconception is that a microdialysis membrane is a detecting device which, in turn leads, incorrectly, to direct comparisons between microdialysis membranes and microvoltammetry indicator detecting devices. Whether or not microdialysis membranes are the same size as voltammetry microelectrodes is irrelevant because the microdialysis membrane is not the detection technology. Microdialysis membranes simply collect perfusate from brain and this perfusate is then analyzed by HPLC.

Dialysis is a technique based on semipermeability of a collection membrane and is not, itself, a detection technique. Existing methods of detecting glutamate by microdialysis followed by HPLC and electrochemical (EC) detection, actually detect a derivative of glutamate rather than glutamate itself. Similarly, microdialysis methods of detecting the neurotransmitter acetylcholine are based on detecting hydrogen peroxide, not acetylcholine itself (Stoecker P W et al., 1990, Selective Electrode Rev. 12:137 160). Moreover, correlation between the derivative of glutamate or H.sub.2O.sub.2 detected and the Cottrell Equation has never been addressed. Therefore, detection of straight chain carbon compounds by the microdialysis membrane method may be questionable. Broderick probes offer an attractive alternative since they may be able to directly detect glutamate or acetylcholine.

Generally, quantitation of neurochemistry is described as a percentage of a few data points, over hours, used as "control" in microdialysis studies. However, Broderick probes are easily calibrated and concentrations are interpolated from calibration curves (Broderick P A et al., 2000, Brain Res. 878:49 63).

One of the advantages of using Broderick probes with microvoltammetry is that microvoltammograms may be obtained from freely moving and behaving living animals and humans. Thus, in some embodiments of the invention another parameter may be monitored and/or recorded. For example, a Broderick probe microvoltammogram may be acquired from a subject while simultaneously monitoring and/or recording the subject's movements (e.g. ambulations and/or fine motor movements). Other examples of parameters that may be monitored and/or recorded include, inter alia, the presence and concentration of a drug, protein, nucleic acid, (e.g. mRNA), carbohydrate, or lipid; consciousness of the subject, cognitive functions, self-administration paradigms, reward-stimulus paradigms, electrophysiological functions, and memory.

The invention provides a variety of methods for identification, diagnosis, and treatment of neuropathologies, neurotoxicities, tumors, and brain and spinal cord injuries using microelectrodes with microvoltammetry. These methods comprise comparing Broderick probe microvoltammograms from at least two different tissues. One these tissues is generally a reference tissue or control. The other is tissue is that being assayed. Preferably, the reference tissue corresponds to the assay tissue with respect to, for example, tissue type, anatomical location, and/or stage of development.

In some embodiments of the invention, the comparison is performed between microvoltammograms taken from the same tissue at different times. In some embodiments, the microvoltammograms compared are taken from the same tissue before and after exposure to a material such as a drug. In some embodiments, a tissue suspected of being diseased is compared with healthy tissue.

Such comparisons may make it possible to diagnose and/or treat a wide variety of diseases or conditions that are associated with abnormal neurotransmitter levels. The invention provides methods comprising exposing at least a cell to a diagnostic challenge or therapeutic treatment, contacting said cell with a Broderick porbe, applying a potential to said Broderick probe; and generating a Broderick probe microvoltammogram. A diagnostic challenge may be designed to elicit a differential response from cells of interest, e.g. diseased cells, from other cells, e.g. healthy cells. A therapeutic treatment may or therapeutic treatment may be known or intended to cure or ameliorate a disease condition. Alternatively, a treatment may be assessed for its capacity to serve as a diagnostic indicator or therapeutic treatment. A diagnostic challenge or a therapeutic treatment may comprise exposing the cell(s) to a material such as a small molecule drug or drug candidate, a defined electrochemical environment (e.g. application of a potential to the cell(s)), exposure to an isotopic or nonisotopic label, activation or repression of a preselected gene, or combinations thereof.

Disorders of basal ganglia, such as athetoid, dystonic diseases, and cancer may be studied with the Broderick probe. An example of an athetoid, dystonic disease is Lesch Nyhan Syndrome (LNS). This recently recognized disease is characterized by severe athetoid and dystonic movements, self-mutilation, and repetitive oral stereotypies. Patients suffering from LNS may have to have their teeth removed to avoid oral stereotypies that cause the patient to devour lips, tongues or fingers. The stereotypies involve DA and 5-HT (Allen SM et al., 1999, Behav. Pharmacol. 10:467 474) and high levels of UA (Patten J, 1980, Neurological Differential Diagnosis, pp. 127 128). Other athetoid and dystonic diseases, such as autism, spinal cord injury, schizophrenia, epilepsy and Parkinson's, are amenable for study with these miniature sensors, even intraoperatively, insofar as epilepsy and Parkinson's are concerned. Several reports indicate that various cancers are also amenable for study with these miniature sensors (Broderick P A, 1989, U.S. Pat. No. 4,883,057; Broderick P A, 1995, U.S. Pat. No. 5,433,710; Broderick P A, 1997, EP 0487647 B1; Broderick P A, 1999, U.S. Pat. No. 5,938,903; Broderick P A, 1999, Hong Kong, HK # 1007350; Broderick P A, 1988, Neurosci. Lett. 95:275 280; Broderick P A, 1989, Brain Res. 495:115 121; Broderick P A, 1990, Electroanalysis 2:241 251; Broderick P A, 2000, Epilepsia 41(Suppl.):91; Broderick P A et al., 2000, Brain Res. 878:49 63).

Much of the difficulty in determining the importance of the alterations in relative concentrations of neurotransmitters and their relationship to epileptogenesis in temporal lobe epilepsy relates to the variability in both the etiology of epilepsy and the location of the epileptogenic zone in epilepsy patients. Few studies have analyzed monoamine concentrations in human epileptic tissue. Those that have studied resected temporal lobe tissue have not distinguished between neocortical temporal lobe epilepsy and mesial temporal lobe epilepsy patients. Neocortical temporal lobe tissue that was part of the ictal onset zone as verified by intracranial EEG recordings of seizures in patients with neocortical temporal lobe epilepsy was examined according to the invention. As previously described (Doyle W K et al., 1997, Epilepsy: A Comprehensive Textbook, pp. 1807 1815), the anterior temporal neocortex in patients with mesial temporal lobe epilepsy is routinely removed at our center to gain access to the mesial temporal structures, providing neocortical tissue controls for our study. While patients with mesial temporal lobe epilepsy may have coexisting neocortical abnormalities like cortical dysplasia (CD), none of the mesial temporal lobe epilepsy patients included in this study had pathologically confirmed CD. Secondary changes such as mild diffuse gliosis were found in the temporal neocortex of mesial temporal lobe epilepsy patients, but we hypothesized that the normal neurochemical profile may still be preserved compared to the actively seizing neocortical tissue analyzed in our neocortical temporal lobe epilepsy patients.

The invention provides devices and methods for diagnosing temporal lobe epilepsy comprising generating a temporally resolved Broderick probe microvoltammogram of a temporal lobe tissue of a subject; and comparing said microvoltammogram to at least one reference Broderick probe microvoltammogram; wherein said reference is a Broderick probe microvoltammogram of the corresponding temporal lobe tissue of another individual. In some embodiments of the invention, the subject's microvoltammogram is compared with one or more reference microvoltammograms from a healthy individual, an individual having mesial temporal lobe epilepsy, an individual having neocortical temporal lobe epilepsy, or combinations thereof.

The invention provides diagnostic devices and methods for brain cancer. In some embodiments the methods comprise: generating a temporally resolved Broderick probe microvoltammetric profile of cancerous cells or tissue; determining from said profile the presence and concentration of at least two markers selected from the group consisting of serotonin, dopamine, ascorbic acid, norepinephrine, .gamma.-aminobutyric acid, glutamate, neurotensin, somatostatin, dynorphin, homovanillic acid, uric acid, tryptophan, tyrosine, nitrous oxide, and nitric oxide; and comparing said marker concentrations to specific threshold values of each of the markers to determine the presence of statistically significant concentration differences, preferably P<0.05; wherein said threshold values are derived from Broderick probe microvoltammetric profile(s) of healthy cells or tissue and said step of comparing said markers distinguishes whether the cancerous cells are present in gray matter or white matter. In other embodiments, the diagnostic methods comprise generating a temporally resolved Broderick probe microvoltammetric profile of a tissue having or at risk of having a tumor; comparing said microvoltammogram to at least one reference Broderick probe microvoltammogram; wherein said reference is a Broderick probe microvoltammogram of corresponding tissue of a healthy individual, cultured cells thereof, corresponding tissue of a an individual having a tumor, cultured cells thereof, or combinations thereof.

The invention also provides diagnostic devices and methods for brain or spinal cord injury. In some embodiments of the invention these methods comprise: generating a temporally resolved Broderick probe microvoltammogram of a tissue of a mammal having or being at risk of developing a brain or spinal cord injury; simultaneously monitoring movement of said mammal; and comparing said microvoltammogram and movement behavior to a reference microvoltammogram of corresponding tissue of a healthy tissue and reference movement behavior of a healthy individual. In addition, the invention provides methods for detecting a site of nerve damage or blockage. These methods may comprise generating a temporally resolved Broderick probe microvoltammogram of a tissue of said mammal; simultaneously monitoring movement of said mammal; and comparing said microvoltammogram and movement behavior to a reference microvoltammogram of corresponding tissue of a healthy tissue and reference movement behavior of a healthy individual.

The invention provides devices and methods for treating temporal lobe epilepsy comprising generating a temporally resolved Broderick probe microvoltammogram of a temporal lobe tissue of a subject having or at risk of developing a temporal lobe epilepsy; comparing said microvoltammogram to at least one reference Broderick probe microvoltammogram; determining the type and extent of temporal lobe resection necessary to achieve a substantially seizure free outcome; and resecting the subject's temporal lobe accordingly. In some embodiments of the invention, the subject's microvoltammogram is compared with one or more reference microvoltammograms from a healthy individual, an individual having mesial temporal lobe epilepsy, an individual having neocortical temporal lobe epilepsy, or combinations thereof.

Brain or spinal cord injuries as well as nerve damage or blockage treatment may include generating Broderick probe microvoltammograms during therapy, i.e. while a pharmacological therapy or kinesitherapy is being administered. Broderick probe microvoltammograms may be acquired continuously during therapy or at intervals. Likewise, cancer treatments may be adapted to include Broderick probe microvoltammetry during therapy. By generating Broderick probe microvoltammograms, it may be possible to monitor tumor size.

The invention contemplates the use of microvoltammetry to assess the neurotoxicity of any material. In some embodiments of the invention, Broderick probe microvoltammograms are acquired from a neural cell or tissue in the presence and absence of the subject material. Materials that may be tested include controlled substances (e.g. opiates, stimulants, depressants, hallucinogens), anti-depressants, anti-epilepsy drugs, and other psychopharmacological substances.

The term "controlled substances" refers to all substances listed in 21 C.F.R. .sctn.1308 even where those referenced only as exceptions. It further includes all salts, geometric and stereoisomers, and derivatives of substances listed therein.

Opiates include, inter alia, alfentanil, alphaprodine, anileridine, apomorphine, bezitramide, carfentanil, cocaine, codeine, 4-cyano-2-dimethylamino-4,4-diphenyl butane, 4-cyano-1-methyl-4-phenylpiperidine pethidine-intermediate-B, dextropropoxyphene, dextrorphan, dihydrocodeine, dihydroetorphine, diphenoxylate, 1-gdiphenylpropane-carboxylic acid pethidine (meperidine), ecgonine, ethyl-4-phenylpiperidine-4-carboxylate pethidine-intermediate-C, ethylmorphine, etorphine hydrochloride, fentanyl, hydrocodone, hydromorphone, isomethadone, levo-alphacetylmethadol, levomethorphan, levorphanol, metazocine, methadone, methadone-intermediate, 2-methyl-3-morpholino-1, 1-methyl-4-phenylpiperidine-4-carboxylic acid, metopon, morphine, moramide-intermediate, nalbuphine, nalmefene, naloxone, naltrexone, opium, oxycodone, oxymorphone, pethidine-intermediate-A, phenanthrene alkaloidsphenazocine, piminodine, racemethorphan, racemorphan, remifentanil, sufentanil, thebaine, and thebaine-derived butorphanol.

Stimulants include substances having a stimulant effect on the central nervous system such as, inter alia, amphetamine, methamphetamine, phenmetrazine, methylphenidate, and salts, isomers, and salts of isomers thereof.

Depressants include substances having a depressant effect on the central nervous system such as, inter alia, amobarbital, glutethimide, pentobarbital, phencyclidine, and secobarbital

Hallucinogens include, inter alia, nabilone.

Anti-depression drugs include, inter alia, citalopram, fluvoxamine, paroxetine, fluoxetine, sertraline, amitriptyline, desipramine, nortriptyline, venlafaxine, phenelzine, tranylcypromine, mirtazepine, nefazodone, trazodone, and bupropion.

Anti-epilepsy drugs include, inter alia, carbamazepine, clorazepate, clopazine, ethosuximide, felbamate, gabapentin, lamotrigine, phenobarbital, phenytoin, primidone, topiramate, and valproic acid.

The neurological side effects including neurotoxicity of any pharmaceutical may be assayed according to the methods of the invention. Neurotoxicity of other substances such as minerals, ions, metals (e.g. heavy metals such as mercury and lead), caffeine, ethanol, nicotine, cannabinoids proteins, lipids, nucleic acids, carbohydrates, glycolipids, and lipoproteins may also be assessed using methods of the invention.
 


Claim 1 of 20 Claims

1. A method of diagnosing and/or monitoring a neurological disorder in a mammal comprising: generating a temporally and/or spatially resolved Broderick probe microvoltammogram of said mammal; determining from said microvoltammogram the presence and concentration of a marker selected from the group consisting of serotonin, dopamine, ascorbic acid, norepinephrine, .gamma.-aminobutyric acid, glutamate, neurotensin, somatostatin, dynorphin, homovanillic acid, uric acid, tryptophan, tyrosine, nitrous oxide, and nitric oxide; and comparing said marker concentration to a threshold value of said respective marker, wherein said threshold value is derived from a Broderick probe microvoltammogram of a mammal that does not have said neurological disorder, wherein the concentration of said marker differs from said threshold value, and wherein said neurological disorder is selected from the group consisting of athetoid, dystonic diseases, epilepsy, Lesch-Nyhan disease, disorders of the basal ganglia, white matter disease, cerebral hemorrhage, head trauma, multiple sclerosis, central nervous system infection, hydrocephalus, and Leukodystrophies.
 

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