Stereotaxic Implantation Research Articles

Implantation of dispersed cells into primate brain.

Although several experimental therapies such as dopaminergic cell implantation in parkinsonian models and intratumoral placement of lymphokine-activated killer cells require intracerebral deposition of dispersed cell suspensions, a successful technique of needle implantation of cells into primate brain has not been demonstrated. The authors have sought to establish a stereotaxic technique to predictably deposit dispersed cells in primate brain. Human lymphocytes were cultured in recombinant interleukin-2, labeled with sodium 51 chromate (51Cr), and stereotaxically injected into the frontal white matter of six anesthetized rhesus monkeys. A 10-microliters aliquot of cell suspension (2 X 10(7) cells/ml) was deposited 16 mm deep to the dura at 5 microliters/min via Hamilton No. 22s or 26s needles. Five control aliquots were counted for each injection. Reflux out of the needle track was absorbed on gauze, and the recovered cells were counted. The animals were sacrificed 1 hour after implantation and the brain was removed and sectioned such that the cortex and white matter along the needle track were separate. The tissue sections were then counted. Recovery was expressed as the percentage of total injected radioactivity (cpm) that was present in each brain section. Two additional injected hemispheres were processed for autoradiography and histological study. Cell recovery in the brain (mean +/- standard deviation) was 87.2% +/- 13.9% (3.3% +/- 4.9% in cortex and 83.9% +/- 15.9% in white matter). The autoradiograms and histological examination showed a dense accumulation of radioactivity (cells) at the target site and minimal radioactivity (cells) in the needle track. Accurate intracerebral deposition of dispersed cells in primates was achieved with the technique described. This knowledge permits reliable stereotaxic implantation of cells into the brains of nonhuman primates and humans for investigation and therapy.

Triiodothyronine exerts direct cell-specific regulation of thyrotropin-releasing hormone gene expression in the hypothalamic paraventricular nucleus.

Thyroid hormone administered systemically exerts negative feedback control of biosynthesis of the TRH pro-hormone in the hypothalamic paraventricular nucleus (PVN), the origin of neurons that regulate anterior pituitary TSH secretion, but not in any other group of TRH-synthesizing neurons in the brain. To determine whether this response is mediated by direct effects on PVN neurons, we studied the effect of unilateral stereotaxic implants of L-T3 into the anterior hypothalamus on the concentration of pro-TRH mRNA and pro-TRH in the PVN of hypothyroid rats. Because hypothalamic-pituitary-thyroid function is also regulated by central catecholamines, we also determined the effect of unilateral ablation of ascending catecholaminergic fibers to one side of the PVN by stereotaxic injection of 6-hydroxydopamine or transection of ascending catecholaminergic pathways. T3-implanted hypothyroid animals showed a marked reduction in pro-TRH mRNA and immunoreactive pro-TRH in medial parvocellular neurons of the PVN on the same side as the implant, but not in contralateral PVN neurons or TRH-synthesizing neurons in other hypothalamic regions. In contrast, hypothyroid animals implanted with pellets of hormonally inactive 3,5-diiodo-L-thyronine showed intense symmetric hybridization and immunoreaction product in both wings of the PVN. Despite marked unilateral reduction in the catecholamine innervation to the PVN, no reduction in pro-TRH mRNA or immunoreactive pro-TRH was observed in the PVN on the affected side compared to that on the unaffected side. These studies demonstrate that negative feedback regulation of thyroid hormone occurs directly on TRH neurons and is restricted only to those in the PVN tuberoinfundibular system.

Neurotensin perfused in hypothalamus of sated or fasted rat: HPLC analysis of release of DA, NE and 5-HT and their metabolites.

This investigation was undertaken in the unrestrained rat to determine the localized effect of neurotensin (NT) on the profile of release and turnover of norepinephrine (NE), dopamine (DA) and serotonin (5-HT) within the hypothalamus. Following stereotaxic implantation of a permanent guide tube, artificial CSF was perfused in the hypothalamus of the freely moving animal by means of push-pull cannulae at a rate of 20 microliters/min and for an interval of 5.0 min. After three 5.0 min control samples were collected, NT in a concentration of 0.1 micrograms/microliter was perfused followed by additional CSF controls. Assay by HPLC-EC of each perfusate showed that when the rat was sated, NT evoked a significant increase in the release of DA and DOPAC from the hypothalamus as well as augmented NE turnover, as reflected by a significant efflux in MHPG. However, when the rat was fasted for 22 hr, the perfusion of NT reduced DA and DOPAC concentrations in the diencephalic perfusate significantly as well as levels of both MHPG and VMA. Under both sated and fasted conditions, NT failed to produce notable changes in the release of 5-HT or its metabolism to 5-HIAA. These findings thus reveal a functional interaction between NT and both of the catecholamine neurotransmitters within hypothalamic neurons, which is clearly dependent upon the nutritional status of the animal.

In vivo electrochemistry with carbon fibre electrodes: Principles and application to neuropharmacology

In vivo voltammetry involves the stereotaxic implantation into the brain of a miniaturised version of the sensitive electrochemical detection system normally used with high-performance liquid chromatography. This article describes the use of working electrodes made from 8 or 30 μm carbon fibres. Their small size makes these electrodes ideally suited for use in small brain nuclei. Chemical and/or electrical pretreatment of these electrodes allows them to be used to monitor the extracellular levels of either neurotransmitter amines such as serotonin and dopamine or their metabolites 5-hydroxyindoleacetic acid and dihydroxyphenylacetic acid.

Stereotaxic implantation of dispersed cell suspensions into brain. A systematic appraisal of cell placement and survival.

The application of several recent advances in cell biology, brain implantation, and cell-mediated tumor immunotherapy requires successful and reproducible placement of viable cell suspensions into brain. Stereotaxic implantation is being used to inject cytotoxic lymphocytes into gliomas and to replace dopaminergic cells in parkinsonian models. Systematic assessment of the factors that influence success in implantation of cell suspensions into solid tissues is needed. A model was developed for investigation of stereotaxic implantation using radiolabeled rat lymphokine-activated killer (LAK) cells. Anesthetized rats received microliter injections of cell suspension into the right caudate nucleus. The injection volume, cell concentration, infusion rate, and needle size were varied systematically. The animals were sacrificed 1 hour after injection; the brain was removed and sectioned, and the radioactivity was counted. Three aliquots of the suspension were injected into counting tubes for control analysis. Recovery of radioactivity was expressed as the percent of mean counts per minute (cpm) in the right frontal lobe/mean cpm in the three control tubes. To assess the viability of implanted cells, the right frontal region was mechanically dissociated in media and centrifuged, and the pellet and supernatant were counted. By using small needles and slow infusion of volumes of 10 microliters or less, 85% to 90% of the radioactivity was recovered in the caudate nucleus. At least half of the implanted cells were viable. Consistent, accurate implantation of dispersed cells into brain over a range of volumes, cell concentrations, infusion rates, and needle sizes was achieved.

Buspirone alters alcohol drinking induced in rats by tetrahydropapaveroline injected into brain monoaminergic pathways

The effect of the novel anxiolytic, buspirone, administered systemically was determined in Sprague-Dawley rats induced to drink ethyl alcohol chronically by repeated microinjections of 25 ng/μl tetrahydropapaveroline HBr (THP) into brain-stem monoaminergic pathways. Self-selection of alcohol in concentrations from 3% to 30% was determined for each rat in a free-choice drinking situation with water available as the alternative fluid. After stereotaxic implantation of guide tubes, THP was microinjected repeatedly into striatal lemniscal and preoptic sites which were found to mediate significant increases in alcohol preference. After the baseline level of intake of a single, maximally preferred alcohol concentration was established, each rat was treated with either saline vehicle or buspirone given intramuscularly b.i.d. in doses of 5.0 mg/kg or 20 mg/kg. Overall, the repeated administration of either dose of buspirone produced a significant decrease in the voluntary alcohol intake of the rats as measured by the proportion measure and absolute g of alcohol ingested. However, the alteration in drinking varied from animal to animal with respect to both magnitude and duration of the anxiolytic's effect. The response to busprione seemed to be dependent in part on the individual site in each animal at which THP had been infused to evoke alcohol intake. Post-mortem histological analysis revealed that buspirone-treated rats reduced alcohol consumption by 83% if THP had been microinjected into substantia nigra; by 60% if given in the nucleus accumbens-preoptic area; and by 34% when injected into the medial lemniscus-zona incerta. These results suggest, therefore, that buspirone can exert a specific effect in attenuating the consumption of alcohol of the rat in a free-choice situation. In relation to the differential actions of the anxiolytic, it is envisaged that on an anatomical basis the antagonism of alcohol drinking may be mediated by a pharmacological alteration of circumscribed pathways associated with dopaminergic or serotonergic neurons.

Book reviewsRadiation Oncology. Vol. 2. Ed. by PhillipsTheodore L. and WaraWilliam, pp. x + 173, 1986 (Raven Press, New York), $75.50. ISBN 0–89004–957–2

This volume is the second in the series originally entitled Radiation Oncology Annual and is intended to be issued at 2–3 year intervals. It is a collection of seven articles, three of which detail treatment techniques, hyperthermia, stereotaxic brain implantation and photodynamic therapy.

Electrical stimulation of the centromedian thalamic nucleus in the treatment of convulsive seizures: a preliminary report.

Five patients with clinical and EEG primary generalized or multifocal uncontrollable seizures underwent stereotaxic implantation of electrodes in both centromedian thalamic nuclei (CM). Each electrode consisted of a semiflexible array of three platinum-iridium wires, isolated except at their tips, which were separated by 4 mm. Bipolar, biphasic rectangular pulses were delivered in trains of 1 min every 5 min, alternating right and left side for sessions 2 h/day. Patients were followed for 3 months with charting of clinical seizures, daily 4-h EEG recordings from scalp and depth for 5 days and weekly thereafter. Baseline and 3-month evaluation of psychological performance through selected Beta R, Wechsler memory scale, visual discrimination, MMPI, and Zung's rated depression scale. Tests were evaluated for significant changes by the parametric student's t test and Mann Whitney nonparametric test. Clinical seizures were significantly reduced by electrical stimulation (ES), as were EEG interictal spikes and EEG slow waves. Psychological performance improved beyond that expected by reduction in seizure activity. ESCM induced a local afterdischarge (AD) that progressively developed in time and intensity, and the beneficial effects outlasted ES for periods of weeks to months, suggesting that a state of hyperexcitability of stimulated tissue, similar to "kindling," was created by chronic ES.

Clinical and dosimetric comparison between I-125 stereotaxic and volume implants for recurrent brain tumors

Clinical and dosimetric comparison between I-125 stereotaxic and volume implants for recurrent brain tumors

Clinical and EEG features of complex partial seizures of temporal lobe origin.

The electrographic and clinical behavioural manifestations of 96 temporal lobe seizures are reviewed from recordings in 19 patients who were submitted to stereotaxic depth electrode implantation in temporal and frontal lobes. Focal onset in hippocampus was recorded in 40% of the seizures. Sixty percent of temporal lobe seizures exhibited a regional seizure onset but in two-thirds of these ictal changes were restricted to amygdaloid and hippocampal structures. Thus, in approximately 80% of seizures, the onset of ictal EEG changes resided in the mesial temporal structures. The main behavioral manifestations observed during seizure discharge restricted to one temporal lobe included warning (67%), motionless stare (24%), automatism (22%), and head-body turning (24%). The predominant ictal behavioural manifestations observed during seizure spread to contralateral temporal and extratemporal structures included warning (3%), motionless stare (36%), automatism (77%), and head-body turning (81%). The direction of head turning did not provide reliable lateralization as to the side of seizure onset.

Gonadotropin secretion following intraventricular norepinephrine infusion into neonatally androgenized female rats.

To determine whether norepinephrine (NE) will stimulate gonadotropin secretion in androgenized female rats, we administered NE (0.3 microM in 2 microliter saline; pH 6.0-6.5) into the third ventricle of females which had been treated neonatally with two different doses of testosterone propionate (TP; 10 or 100 micrograms) on day 5 of life. This treatment rendered these animals anovulatory as evidenced by polyfollicular ovaries and a persistent vaginal estrus condition which occurred by 100 days of age. Those females which were determined to be anovulatory were ovariectomized 4 weeks prior to the stereotaxic implantation of an intracerebroventricular (ICV) cannula. Ten to twelve days following cannula placement, animals were primed with either estradiol benzoate (EB; 30 micrograms) or EB and progesterone (P; 5 mg). Two days following steroid priming, NE was infused into the third ventricle at 15.00 h and blood samples were taken via an intra-atrial catheter at 15-min intervals for 2 h before and 2 h after administration. In both TP-treated groups, NE caused rapid and significant (p less than 0.01) elevations in plasma luteinizing hormone (LH) concentrations when animals were primed with EB as compared with control females given acid saline infusions (pH 6.0-6.5). Similar responses to NE were seen after additional priming with P at the same time as EB. Follicle-stimulating hormone (FSH) values did not increase following NE or saline infusion to any of the groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of hematoporphyrin derivative photoradiation therapy on survival in the rat 9L gliosarcoma brain-tumor model.

Intracerebral tumors were produced in 99 rats by stereotaxic implantation of 9L gliosarcoma brain-tumor cells. Hematoporphyrin derivative (HPD), 10 or 20 mg/kg, was administered as an intravenous bolus 24 or 48 hours before irradiation of the tumor region with light from an argon pumped-dye laser (632 nm). Laser light, at a dose of 30, 60, or 200 joules/sq cm, was delivered through a craniectomy 10 or 13 days after tumor implantation. Survival times were significantly prolonged in rats exposed to laser light at a dose of 200 joules/sq cm 24 hours after administration of HPD, 20 mg/kg.

Release of norepinephrine from the rat's hypothalamus perfused with alcohol: Relation to body temperature

The activity of norepinephrine (NE), within the thermosensitive region of the anterior hypothalamic, pre-optic area (AH/POA) of the rat was examined in relation to changes in core temperature produced by ethyl alcohol. Following stereotaxic implantation of push-pull guide tubes, a specific site in the AH/POA, reactive or non-reactive to NE, was labeled with 1.0 μl of [ 3H]-NE. Alcohol in a concentration of 2.75% or 5.5% was then perfused locally at the same site by push-pull cannulae or administered peripherally in a dose of 2.0 g/kg. In control experiments, artificial CSF was perfused alone. The perfusion of alcohol enhanced or delayed the release of [ 3H]-NE in AH/POA or failed to alter the efflux of the catecholamine, with the specific response dependent principally on the: (1) anatomical site of hypothalamic perfusion, (2) concentration of alcohol, and (3) interval of perfusion itself. During the perfusion of alcohol within a very circumscribed region in the AH/POA, vasodilatation, as reflected by an increase in skin temperature, and a hypothermia of short latency, occurred. The change in core temperature was usually accompanied by a delay in the efflux of [ 3H]-NE. After the peripheral administration of 2.0 g/kg alcohol, an alteration in NE efflux from the AH/POA was also induced during the course of a hypothermic response accompanied by vasodilatation. These results suggest that alcohol exerts a direct central effect on nerve cells comprising the thermoregulatory mechanism located within the hypothalamus. Further, the well-known thermolytic effect of alcohol could be mediated in part by noradrenergic synapses within AH/POA, by means of their phasic release of NE.

Temperature measurements in high thermal gradients: I. The effects of conduction

Two temperature probes (a fluoroptic sensor and a metallic thermistor), which are both suitable for stereotaxic implantation, were used in comparative thermometry studies during interstitial microwave heating of the brain in vivo. Thermal distributions having large temperature gradients (5–10°C/cm) were routinely observed. The temperature differentials (ΔT) between the 2 probes were position dependent within the thermal field. The maximum difference in temperatures measured, using the 2 probes along identical tracks without a catheter, ranged between 0.5°C and 1.8°C. Near the brain/air surface, the thermistor measured lower temperatures than the optical probe; however, medial to the antenna, the thermistor temperatures were higher than the optic sensor. The measured temperature discrepancies are the result of smearing due to thermal conduction along the axial length of the metallic thermistor probe. These effects are significantly accentuated when the temperature probes are tracked in catheters. Experiments performed in a nonperfused phantom, heated with the interstitial microwave antenna, demonstrated similar conductive effects. Studies in a nonelectromagnetic environment (flow cell-thermal step gradient) additionally confirmed that thermal conductive artifacts were the major source of temperature error.

Computerized Tomography-guided Stereotaxic Implantation of Deep Brainstimulating Electrodes

High-resolution computerized tomography (CT) images can detect deep brain commissures, internal capsules and pathological lesions. The intercommissural plane can be determined by a reconstructed sagittal CT image of deep midline structures. The individual CT atlas of the brain of a patient permits precise target identification, and eliminates the need for contrast ventriculography. Three patients, who underwent successful CT-guided stereotaxic implantation of deep brainstimulating electrodes using a Brown-Roberts-Wells apparatus for the control of chronic pain, are reported. The target of two patients who had intractable pain caused by cancer was the thalamic relay nucleus. As the stimulation was effective in controlling the pain, the amounts of analgesics could be reduced. The target of one patient who had thalamic pain was the internal capsule and the pain relief has been excellent. Intraoperative electrophysiological assessments were available to confirm the correct target. Stereotaxic implantation of deep brain electrodes may be the best indication of CT-guided functional neurosurgery. Since this method is easy and accurate, its application to poor risk patient is feasible.

Neuronal and neurovascular integration following transplantation of the fetal hypothalamus into the third cerebral ventricle of adult brattleboro rats. Neurological transplants: I

This investigation has confronted some very basic questions of neurobiology and specifically deals with the neurovascular and neuroanatomical interactions that occur between graft and host following neural transplantation. Host Long-Evans rats with chronic autosomal diabetes insipidus (DI) received stereotaxic implants of normal 17 day post-coitus fetal hypothalamic fragments from the rostral (anterior) hypothalamus of normal Long-Evans pups. Following stereotaxic surgery DI hosts were killed 60 or 90 days later and their brain prepared for correlative microangiography-immunocytochemistry coupled with transmission electron microscopy. Explants were rapidly invaded by host vessels from several routes. 1. (1) Vessels appeared to arise from portal capillaries in the underlying median eminence and 2. (2) from adjacent vessels from the paraventricular nucleus and surrounding endocrine hypothalamus and 3. (3) possibly from intrinsic vessels of the graft. The former remained fenestrated and established bonafide neurovascular zones in the ventral regions and in actively growing explants. Small clusters of arginine vasopressin-positive fibers and neurophysin positive neurons were noted throughout the parenchyma of explants. Despite the presence of neurosecretory neurons and neurovascular (neurohemal) zones, none of the host rats exhibited a physiological return to normal parameters of water balance. However the active growth and development of explants in the third cerebral ventricle of DI host rats coupled with emergence of neurovascular zones lends support to a potential model for analyzing the development of anatomical substrates for the central delivery of neuropeptide hormones.

Ontogeny of cortical and subcortical electroencephalographic events in unrestrained neonatal and infant rats

We have devised a method of stereotaxic implantation of electrodes in cortical and subcortical sites in neonatal and infant rats and subsequently recording from these animals as they move about freely. We studied the development of the EEG recorded from these cortical and subcortical sites in animals ranging in age from 1 to 84 days and analyzed the EEGs both visually and by power spectral analysis via a fast Fourier transform algorithm. The animals survived the procedure with less than 5% overall mortality. Although of very low voltage and very irregular in appearance, EEG activity was present in all sites at 1 day of age. There was gradual increase in synchrony, organization, and voltage until 12 to 14 days when an adult pattern appeared.

Transplantation of fetal rat islets into the cerebral ventricles of alloxan-diabetic rats. Amelioration of diabetes by syngeneic but not allogeneic islets.

Syngeneic fetal rat islets were isolated and transplanted into alloxan-diabetic inbred male Lewis rats. The effect of transplantation of islets into the cerebral ventricles on the diabetic state of the recipients was compared with that of the conventional transplantation of islets homeotypically into the liver via the portal vein. Fourteen of fourteen rats surviving after stereotaxic implantation of islets into the ventricles returned to normoglycemia; normoglycemia has been maintained for up to 34 wk. Glucose tolerance tests revealed an improved, although not completely normalized, pattern. Histologic examination of the brains of these recipients revealed clusters of intact islets in the ventricle. These data provided a physiologic basis for further investigation of the immunologically privileged nature of the intraventricular space as a site for implantation of allogeneic pancreatic islets. Islets from Wistar-Furth rats (major histocompatibility difference) or Fischer 344 rats (minor histocompatibility difference) were transplanted into the ventricles of alloxan-diabetic Lewis rats. There were only small and unsustained changes in body weight and blood and urine glucose of any of the rats receiving the allogeneic islets. Histologic examination of the ventricles of these rats 3 wk after transplantation revealed only glial scar tissue. These data suggest that the cerebral ventricles cannot serve as a privileged site for islet transplantation, at least using the type of islet preparation employed in these experiments.

Glucagon release after stimulation of the lateral hypothalamic area in rats: predominant beta-adrenergic transmission and involvement of endorphin pathways.

Catheterization of the portal vein and stereotaxic implantation of electrodes in the lateral hypothalamic area (LHA) were performed in normal rats after thiopental anesthesia. Immunoreactive glucagon (IRG) and insulin (IRI), glucose, catecholamines, and beta-endorphin were monitored in portal and peripheral plasma, before and during electrical stimulation of the LHA. The influences on glucose and hormone concentrations of propranolol, phentolamine, atropine, and naloxone infusions were also investigated in similar rats. A basal portoperipheral concentration gradient was found for IRG, IRI, and catecholamines, but not for beta-endorphin. The LHA stimulation induced a significant rise in portal catecholamine, IRG, and glucose concentrations; IRI remained unchanged; the portoperipheral catecholamine gradient was augmented. These alterations were not observed after bilateral splanchnicectomy. Propranolol infusion abolished the LHA-dependent IRG and glucose rises. Naloxone reduced the IRG rise significantly. Phentolamine and atropine did not modify the LHA-induced reactions. These results suggest that the glucagon release which follows LHA electrical stimulation depends mainly on beta-adrenergic transmission by the splanchnic nerves. Opioid peptide receptors may modulate this effect.

Effects of histamine and some biogenic amines on the hippocampal after-discharge in rats.

The stereotaxic implantation of chronic electrodes for EEG recording was done into the frontal cortex, septal region and amygdala, and simultaneous insertion of a cannula into the lateral ventricle was conducted. Indifferent electrode was placed in the occipital part of the cranium, and bipolar electrode for electrical stimulation was also implanted into the hippocampus. At least 10 days were elapsed before the initiation of experiment. The animals were placed in a plastic observation chamber, and EEG was recorded with monopolar electrodes. The after-discharge (AD) was provoked by electrical stimulation (3 msec duration, 60 Hz) to the hippocampus for 2 sec. Intraventricular application of 100 μg histamine (Hi) decreased the AD duration determined at 5 and 30 min later, significantly. However, at a dose of 500 μg, Hi caused significant increase of the AD duration even after 60 and 120 min. 2-MethylHi (100 μg) depressed AD; however, 4-MethylHi (200 μg) extended the AD duration. Diphenhydramine (1 mg/kg, i.v.) and pyrilamine (5 mg/kg, i.v.) prevented Hi-induced suppression of AD, but Cimetidine (2 mg/kg, i.v.) and ranitidine (2 mg/kg, i.v.) did not affect the AD. Although noradrenaline (200 μg), serotonin (100 μg) and dopamine (100 μg) did not affect the AD duration, acetylcholine (100 μg) induced significant prolongation. GABA (200 μg), glycine (200 μg) and taurine (200 μg) did not affect hippocampal AD. From these results, it has been assumed that the suppression of AD induced by Hi can not be attributed to the effect mediated by these amines and amino acids; and suppressive effect of Hi on AD can be associated with H1 receptor, while prolongation of AD by Hi may be related with H2 receptor.