Adaptation of tape removal test for measurement of sensitivity in perineal area of rat.

Regeneration after spinal cord injury is a goal of many studies. Although the most obvious target is to recover motor function, restoration of sensation can also improve the quality of life after spinal cord injury. For many patients, recovery of sensation in the perineal and genital area is a high priority. Currently there is no experimental test in rodents for measuring changes in sensation in the perineal and genital area after spinal cord injury. The aim of our study was to develop a behavioural test for measuring the sensitivity of the perineal and genital area in rats. We have modified the tape removal test used routinely to test sensorimotor deficits after stroke and spinal cord injury to test the perineal area with several variations. A small piece of tape (approximately 1 cm2) was attached to the perineal area. Time to first contact and to the removal of the tape was measured. Each rat was trained for 5 consecutive days and then tested weekly. We compared different rat strains (Wistar, Sprague-Dawley, Long-Evans and Lewis), both genders, shaving and non-shaving and different types of tape. We found that the test was suitable for all tested strains, however, Lewis rats achieved the lowest contact times, but this difference was significant only for the first few days of learning the task. There were no significant differences between gender and different types of tape or shaving. After training the animals underwent dorsal column lesion at T10 and were tested at day 3, 8, 14 and 21. The test detected a sensory deficit, the average time across all animals to sense the stimulus increased from 1'32 up to 3'20. There was a strong relationship between lesion size and tape detection time, and only lesions that extended laterally to the dorsal root entry zone produced significant sensory deficits. Other standard behavioural tests (BBB, von Frey, ladder and Plantar test) were performed in the same animals. There was a correlation between lesion size and deficit for the ladder and BBB tests, but not for the von Frey and Plantar tests. We conclude that the tape removal test is suitable for testing perineal sensation in rats, can be used in different strains and is appropriate for monitoring changes in sensation after spinal cord injury.

Short Review: Investigating ARSACS: models for understanding cerebellar degeneration.

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is an early-onset neurodegenerative disease that includes progressive cerebellar dysfunction. ARSACS is caused by an autosomal recessive loss-of-function mutation in the SACS gene, which encodes for SACSIN. Although animal models are still necessary to investigate the role of SACSIN in the pathology of this disease, more reliable human cellular models need to be generated to better understand the cerebellar pathophysiology of ARSACS. The discovery of human induced pluripotent stem cells (hiPSC) has permitted the derivation of patient-specific cells. These cells have an unlimited self-renewing capacity and the ability to differentiate into different neural cell types, allowing studies of disease mechanism, drug discovery and cell replacement therapies. In this study, we discuss how the hiPSC-derived cerebellar organoid culture offers novel strategies for targeting the pathogenic mutations related to ARSACS. We also highlight the advantages and challenges of this 3D cellular model, as well as the questions that still remain unanswered.

Stem cells regenerative properties on new rat spinal fusion model.

Stem cells biology is one of the most frequent topic of physiological research of today. Spinal fusion represents common bone biology challenge. It is the indicator of osteoinduction and new bone formation on ectopic model. The purpose of this study was to establish a simple model of spinal fusion based on a rat model including verification of the possible use of titanium microplates with hydroxyapatite scaffold combined with human bone marrow-derived mesenchymal stem cells (MSCs). Spinous processes of two adjacent vertebrae were fixed in 15 Wistar rats. The space between bony vertebral arches and spinous processes was either filled with augmentation material only and covered with a resorbable collagen membrane (Group 1), or filled with augmentation material loaded with 5 × 106 MSCs and covered with a resorbable collagen membrane (Group 2). The rats were sacrificed 8 weeks after the surgery. Histology, histomorphometry and micro-CT were performed. The new model of interspinous fusion was safe, easy, inexpensive, with zero mortality. We did not detect any substantial pathological changes or tumor formation after graft implantation. We observed a nonsignificant effect on the formation of new bone tissue between Group 1 and Group 2. In the group with MSCs (Group 2) we described minor inflamatory response which indicates the imunomodulational and antiinflamatory role of MSCs. In conclusion, this new model proved to be easy to use in small animals like rats.

Experimental reconstruction of the injured spinal cord.

Injury to the spinal cord, with its pathological sequelae, results in a permanent neurological deficit. With currently available tools at hand, there is very little that clinicians can do to treat such a condition with the view of helping patients with spinal cord injury (SCI). On the other hand, in the last 20 years experimental research has brought new insights into the pathophysiology of spinal cord injury; we can divide the time course into 3 phases: primary injury (the time of traumatic impact and the period immediately afterwards), the secondary phase (cell death, inflammation, ischemia), and the chronic phase (scarring, demyelination, cyst formation). Increased knowledge about the pathophysiology of SCI can stimulate the development of new therapeutic modalities and approaches, which may be feasible in the future in clinical practice. Some of the most promising experimental therapies include: neurotrophic factors, enzymes and antibodies against inhibitory molecules (such as Nogo), activated macrophages, stem cells and bridging scaffolds. Their common goal is to reconstitute the damaged tissue in order to recover the lost function. In the current review, we focus on some of the recent developments in experimental SCI research.

[Comparison of chondrogenic differentiation of adipose tissue-derived mesenchymal stem cells with cultured chondrocytes and bone marrow mesenchymal stem cells]. / Výsledky srovnání chondrogenní diferenciace mezenchymálních kmenových bunek získaných z tukové tkáne s kultivovanými chondrocyty a mezenchymálními kmenovými bunkami z kostní drene

PURPOSE OF THE STUDY: Many congenital and acquired disorders as well as sequelae of injury are associated with articular cartilage degeneration, which adversely affects the patient's quality of life. The currently used cell therapy with cultured chondrocytes has its disadvantages due to a process of de-differentiation of chondrocytes during cultivation. We believe that the mesenchymal stem cell therapy offers a new treatment options. MATERIAL AND METHODS: The adult mesenchymal stem cells (MSCs) for chondrocyte differentiation are usually obtained from bone marrow mesenchymal stem cells (BMSCs). In this study these cells were compared with mesenchymal stem cells derived from adipose tissue (AMSCs). The aim of the study was to verify the ability of human BMSCs and AMSCs to differentiate into chondrocytes in vitro in the presence or absence of transforming growth factor beta (TGF-ß1). Human BMSCs and AMSCs were collected from healthy donors during orthopaedic surgeries, in vitro cultured in three passages to obtain the required quantity of cells. A pellet culture system was used for chondrocyte differentiation. RESULTS: At 21 days of cultivation, cell aggregates grown in the chondrogenic medium were larger than those cultured in the control medium. Both the BMSCs and AMSCs pellet cultures showed spontaneous chondrogenesis. Histological staining with haematoxylin and eosin and Masson's trichrome stains, as well as immunohistochemical staining to detect type II collagen revealed no apparent differences between the pellet cultures with TGF-ß1 presence and those without it. The real-time RT-PCR detected expression of the type II collagen gene in all tested cultures. In the BMSCs pellet culture only, TGF-ß1 presence resulted in a decrease in type I collagen mRNA levels and in an increase in type II collagen mRNA values. DISCUSSION: Our results showed an in vitro chondrogenic potential of mature human mesenchymal stem cells derived from both bone marrow and adipose tissue. In agreement with the relevant literature data, we suggest that both cell types have an equal prospect for use in cartilage tissue engineering.

Core-shell La(1-x)Sr(x)MnO3 nanoparticles as colloidal mediators for magnetic fluid hyperthermia.

Core-shell nanoparticles consisting of La(0.75)Sr(0.25)MnO(3) cores covered by silica were synthesized by a procedure consisting of several steps, including the sol-gel method in the presence of citric acid and ethylene glycol, thermal and mechanical treatment, encapsulation employing tetraethoxysilane and final separation by centrifugation in order to get the required size fraction. Morphological studies revealed well-separated particles that form a stable water suspension. Magnetic studies include magnetization measurements and investigation of the ferromagnetic-superparamagnetic-paramagnetic transition. Magnetic heating experiments in 'calorimetric mode' were used to determine the heating efficiency of the particles in water suspension and further employed for biological studies of extracellular and intracellular effects analysed by tests of viability.

Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part 6: 3D hydrogels with positive and negative surface charges and polyelectrolyte complexes in spinal cord injury repair.

Macroporous hydrogels are artificial biomaterials commonly used in tissue engineering, including central nervous system (CNS) repair. Their physical properties may be modified to improve their adhesion properties and promote tissue regeneration. We implanted four types of hydrogels based on 2-hydroxyethyl methacrylate (HEMA) with different surface charges inside a spinal cord hemisection cavity at the Th8 level in rats. The spinal cords were processed 1 and 6 months after implantation and histologically evaluated. Connective tissue deposition was most abundant in the hydrogels with positively-charged functional groups. Axonal regeneration was promoted in hydrogels carrying charged functional groups; hydrogels with positively charged functional groups showed increased axonal ingrowth into the central parts of the implant. Few astrocytes grew into the hydrogels. Our study shows that HEMA-based hydrogels carrying charged functional groups improve axonal ingrowth inside the implants compared to implants without any charge. Further, positively charged functional groups promote connective tissue infiltration and extended axonal regeneration inside a hydrogel bridge.

Rapid but not slow spinal cord compression elicits neurogenic pulmonary edema in the rat.

The development of neurogenic pulmonary edema (NPE) can be elicited by an immediate epidural balloon compression of the thoracic spinal cord. To evaluate whether a slower balloon inflation could prevent NPE development, we examined the extent of NPE in animals lesioned with a rapid (5 microl - 5 microl - 5 microl) or slow rate (3 microl - 2 microl - 2 microl - 2 microl - 2 microl - 2 microl - 2 microl) of balloon inflation. These groups were compared with the NPE model (immediate inflation to 15 microl) and with healthy controls. Slow balloon inflation prevented NPE development, whereas the pulmonary index and histology revealed a massive pulmonary edema in the group with a rapid rate of balloon inflation. Pulmonary edema was preceded by a considerable decrease in heart rate during the inflation procedure. Moreover, rapid inflation of balloon in spinal channel to either 5 microl or 10 microl did not cause NPE. Thus, a slow rate of balloon inflation in the thoracic epidural space prevents the development of neurogenic pulmonary edema, most likely due to the better adaptation of the organism to acute circulatory changes (rapid elevation of systemic blood pressure accompanied by profound heart rate reduction) during the longer balloon inflation period. It should be noted that spinal cord transection at the same level did not cause neurogenic pulmonary edema.

Biocompatible hydrogels in spinal cord injury repair.

Spinal cord injury results in a permanent neurological deficit due to tissue damage. Such a lesion is a barrier for "communication" between the brain and peripheral tissues, effectors as well as receptors. One of the primary goals of tissue engineering is to bridge the spinal cord injury and re-establish the damaged connections. Hydrogels are biocompatible implants used in spinal cord injury repair. They can create a permissive environment and bridge the lesion cavities by providing a scaffold for the regeneration of neurons and their axons, glia and other tissue elements. The advantage of using artificial materials is the possibility to modify their physical and chemical properties in order to develop the best implant suitable for spinal cord injury repair. As a result, several types of hydrogels have been tested in experimental studies so far. We review our work that has been done during the last 5 years with various types of hydrogels and their applications in experimental spinal cord injury repair.

Mechanisms of neurogenic pulmonary edema development.

Neurogenic pulmonary edema is a life-threatening complication, known for almost 100 years, but its etiopathogenesis is still not completely understood. This review summarizes current knowledge about the etiology and pathophysiology of neurogenic pulmonary edema. The roles of systemic sympathetic discharge, central nervous system trigger zones, intracranial pressure, inflammation and anesthesia in the etiopathogenesis of neurogenic pulmonary edema are considered in detail. The management of the patient and experimental models of neurogenic pulmonary edema are also discussed.

Low degree of anesthesia increases the risk of neurogenic pulmonary edema development.

Neurogenic pulmonary edema is an acute life-threatening complication following central nervous system injury. The exact pathogenic mechanism leading to its development is still unclear. We introduce a new hypothesis that high levels of anesthesia might protect the organism against the development of neurogenic pulmonary edema due to a more pronounced inhibition of the hypothalamic, brainstem and spinal vasoactive sympathetic centers. On the basis of a more pronounced neuronal inhibition of the vasoactive centers, a severe sympathetic discharge does not occur and neurogenic pulmonary edema does not develop. In contrast, an insufficient anesthesia level is not able to inhibit the sympathetic nervous system during an injury of the central nervous system and thus neurogenic pulmonary edema develops. During experiments with central nervous system injury, low-anesthesia-induced neurogenic pulmonary edema might negatively influence the overall recovery of the animal. More importantly, during a neurosurgical intervention, insufficient anesthesia might similarly lead to neurogenic pulmonary edema development in operated patients. Our hypothesis indicates the necessity of precisely monitoring of the level anesthesia during experimental manipulations of the central nervous system in animals or neurosurgical interventions in humans.

Migration, fate and in vivo imaging of adult stem cells in the CNS.

Adult stem cells have been intensively studied for their potential use in cell therapies for neurodegenerative diseases, ischemia and traumatic injuries. One of the most promising cell sources for autologous cell transplantation is bone marrow, containing a heterogenous cell population that can be roughly divided into hematopoietic stem and progenitor cells and mesenchymal stem cells (MSCs). MSCs are multipotent progenitor cells that, in the case of severe tissue ischemia or damage, can be attracted to the lesion site, where they can secrete bioactive molecules, either naturally or through genetic engineering. They can also serve as vehicles for delivering therapeutic agents. Mobilized from the marrow, sorted or expanded in culture, MSCs can be delivered to the damaged site by direct or systemic application. In addition, MSCs can be labeled with superparamagnetic nanoparticles that allow in vivo cell imaging. Magnetic resonance imaging (MRI) is thus a suitable method for in vivo cell tracking of transplanted cells in the host organism. This review will focus on cell labeling for MRI and the use of MSCs in experimental and clinical studies for the treatment of brain and spinal cord injuries.

Macroporous hydrogels based on 2-hydroxyethyl methacrylate. Part 4: growth of rat bone marrow stromal cells in three-dimensional hydrogels with positive and negative surface charges and in polyelectrolyte complexes.

The growth of bone marrow stromal cells was assessed in vitro in macroporous hydrogels based on 2-hydro- xyethyl methacrylate (HEMA) copolymers with different electric charges. Copolymers of HEMA with sodium methacrylate (MA(-)) carried a negative electric charge, copolymers of HEMA with [2-(methacryloyloxy)ethyl] trimethylammonium chloride (MOETA(-)) carried a positive electric charge and terpolymers of HEMA, MA(-) and MOETA(+) carried both, positive and negative electric charges. The charges in the polyelectrolyte complexes were shielded by counter-ions. The hydrogels had similar porosities, based on a comparison of their diffusion parameters for small cations as measured by the real-time tetramethylammonium iontophoretic method of diffusion analysis. The cell growth was studied in the peripheral and central regions of the hydrogels at 2 hours and 2, 7, 14 and 28 days after cell seeding. Image analysis revealed the highest cellular density in the HEMA-MOETA(+) copolymers; most of the cells were present in the peripheral region of the hydrogels. A lower density of cells but no difference between the peripheral and central regions was observed in the HEMA-MA(-) copolymers and in polyelectrolyte complexes. This study showed that positively charged functional groups promote the adhesion of cells.

Green fluorescent protein bone marrow cells express hematopoietic and neural antigens in culture and migrate within the neonatal rat brain.

Finding a reliable source of alternative neural stem cells for treatment of various diseases and injuries affecting the central nervous system is a challenge. Numerous studies have shown that hematopoietic and nonhematopoietic progenitors derived from bone marrow (BM) under specific conditions are able to differentiate into cells of all three germ layers. Recently, it was reported that cultured, unfractionated (whole) adult BM cells form nestin-positive spheres that can later initiate neural differentiation (Kabos et al., 2002). The identity of the subpopulation of BM cells that contributes to neural differentiation remains unknown. We therefore analyzed the hematopoietic and neural features of cultured, unfractionated BM cells derived from a transgenic mouse that expresses green fluorescent protein (GFP) in all tissues. We also transplanted the BM cells into the subventricular zone (SVZ), a region known to support postnatal neurogenesis. After injection of BM cells into the neurogenic SVZ in neonatal rats, we found surviving GFP+ BM cells close to the injection site and in various brain regions, including corpus callosum and subcortical white matter. Many of the grafted cells were detected within the rostral migratory stream (RMS), moving toward the olfactory bulb (OB), and some cells reached the subependymal zone of the OB. Our in vitro experiments revealed that murine GFP+ BM cells retained their proliferation and differentiation potential and predominantly preserved their hematopoietic identity (CD45, CD90, CD133), although a few expressed neural antigens (nestin, glial fibrillary acdiic protein, TuJ1).

Glutamate, NMDA, and AMPA induced changes in extracellular space volume and tortuosity in the rat spinal cord.

Glutamate release, particularly in pathologic conditions, may result in cellular swelling. The authors studied the effects of glutamate, N-methyl-D-aspartate (NMDA), and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) on extracellular pH (pH(e)), extracellular potassium concentration ([K(+)](e)), and changes in extracellular space (ECS) diffusion parameters (volume fraction alpha, tortuosity lambda) resulting from cellular swelling. In the isolated spinal cord of 4-to 12-day-old rats, the application of glutamate receptor agonists induced an increase in [K(+)](e), alkaline-acid shifts, a substantial decrease in alpha, and an increase in lambda. After washout of the glutamate receptor agonists, alpha either returned to or overshot normal values, whereas lambda remained elevated. Pretreatment with 20 mmol/L Mg(++), MK801, or CNQX blocked the changes in diffusion parameters, [K(+)](e) and pH(e) evoked by NMDA or AMPA. However, the changes in diffusion parameters also were blocked in Ca(2+)-free solution, which had no effect on the [K(+)](e) increase or acid shift. The authors conclude that increased glutamate release may produce a large, sustained and [Ca(2+)](e)-dependent decrease in alpha and increase in lambda. Repetitive stimulation and pathologic states resulting in glutamate release therefore may lead to changes in ECS volume and tortuosity, affecting volume transmission and enhancing glutamate neurotoxicity and neuronal damage.

K+ and pH homeostasis in the developing rat spinal cord is impaired by early postnatal X-irradiation.

Activity-related transient changes in extracellular K+ concentration ([K+]e) and pH (pHe) were studied by means of ion-selective microelectrodes in neonatal rat spinal cords isolated from pups 2-14 days of age. Pups 1 to 2 days old were X-irradiated to impair gliogenesis and spinal cords were isolated 2-13 days postirradiation (PI). In 2- to 14-day-old pups PI stimulation produced ionic changes that were the same as those in 3- to 6-day-old control (non-irradiated) pups; e.g. the [K+]e increased by 4.03 +/- 0.24 mM (mean +/- S.E.M., n = 30) at a stimulation frequency of 10 Hz and this was accompanied by an alkaline shift of 0.048 +/- 0.004 pH units (mean +/- S.E.M., n = 32) pH units. By contrast, stimulation in non-irradiated 10- to 14-day-old pups produced smaller [K+]e changes, of 1.95 +/- 0.12 mM (mean +/- S.E.M., n = 30), and an acid shift of 0.035 +/- 0.003 pH units which was usually preceded by a scarcely discernible initial alkaline shift, as is also the case in adult rats. Our results show that the decrease in [K+]e ceiling level and the development of the acid shift in pHe are blocked by X-irradiation. Concomitantly, typical continuous development of GFAP-positive reaction was disrupted and densely stained astrocytes in gray matter of 10- to 14-day-old pups PI revealed astrogliosis. In control 3- to 6-day-old pups and in pups PI the stimulation-evoked alkaline, but not the acid, shift was blocked by Mg2+ and picrotoxin (10(-6) M). The acid shift was blocked, and the alkaline shift enhanced, by acetazolamide, Ba2+, amiloride and SITS. Application of GABA evoked an alkaline shift in the pHe baseline which was blocked by picrotoxin and in HEPES-buffered solution. By contrast, the stimulus-evoked alkaline shifts were enhanced in HEPES-buffered solutions. The results suggest a dual mechanism of the stimulus-evoked alkaline shifts. Firstly, the activation of GABA-gated anion (Cl-) channels induces a passive net efflux of bicarbonate, which may lead to a fall in neuronal intracellular pH and to a rise in the pHe. Secondly, bicarbonate independent alkaline shifts may arise from synaptic activity resulting in a flux of acid equivalents.

Extracellular K+, pH, and volume changes in spinal cord of adult rats and during postnatal development.

Activity-related transient changes in extracellular K+ concentration ([K+]c), extracellular pH (pHc), and extracellular volume (EC volume) were studied by means of ion-selective microelectrodes in the adult rat spinal cord in vivo and in neonatal rat spinal cords isolated from pups 3-14 days of age. Repetitive electrical nerve stimulation (10-100 Hz) in adults elicited increases in [K+]c by about 2.0-3.5 mM, followed by a poststimulation K+ undershoot and triphasic alkaline-acid-alkaline changes in pHc. In 3- to 6-day-old pups, the [K+]c increased by as much as 6.5 mM at a stimulation frequency of 10 Hz, and this was accompanied by an alkaline shift. Increases in [K+]c as large as 1.3-2.5 mM accompanied by an alkaline shift were evoked by a single electrical stimulus. Stimulation in 10- to 13-day-old pups produced smaller [K+]c change and an acid shift, which was preceded by a small initial alkaline shift, as in adult rats. We conclude that glial cells buffer the activity-related [K+]c increase and alkaline pHc shifts. Mg2+ blocked the alkaline but not the acid shift. Acetazolamide had no effect on the alkaline shift but blocked the acid shift. The alkaline shift was enhanced and the acid shift blocked by Ba2+, amiloride, 4-acetamido-4'-isothiocyanotostilbene-2,2'-disulfonic acid (SITS), and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS). Activity-related acid shifts therefore have a complex mechanism, which includes Na+H+ exchange, Cl-/HCO3- exchange, or Na+/Cl-/H+/HCO3- antiport, Na(+)-HCO3- cotransport, and H+ efflux through voltage-sensitive H+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of glia in K+ and pH homeostasis in the neonatal rat spinal cord.

Stimulation-evoked transient changes in extracellular potassium ([K+]e) and pH (pHe) were studied in the neonatal rat spinal cords isolated from 3-13-day-old pups. In unstimulated pups the [K+]e baseline was elevated and pHe was more acid than that in Ringer's solution (3.5 mM K+, pH 7.3-7.35). The [K+]e and pHe in 3-6-day-old pups was 3.91 +/- 0.12 mM and pHe 7.19 +/- 0.01, respectively, while in 10-13-day-old pups it was 4.35 +/- 0.15 mM and 7.11 +/- 0.01, respectively. The [K+]e changes evoked in the dorsal horn by a single electrical stimulus were as large as 1.5-2.5 mM. Such changes in [K+]e are evoked in the adult rat spinal cord with stimulation at a frequency of 10-30 Hz. The maximal changes of 2.1-6.5 mM were found at a stimulation frequency of 10 Hz in 3-6-day-old animals. In older animals the [K+]e changes progressively decreased. The poststimulation K(+)-undershoot was found after a single stimulus as well as after repetitive stimulation. In 3-8-day-old pups, the stimulation evoked an alkaline shift, which was followed by a smaller poststimulation acid shift when the stimulation was discontinued. In pups 3-4-days-old the stimulation evoked the greatest alkaline shifts, i.e., by as much as 0.05 pH units after a single pulse and by about 0.1 pH units during stimulation at a frequency of 10 Hz. In 5-8-day-old pups, the alkaline shift became smaller and the poststimulation acid shift increased.(ABSTRACT TRUNCATED AT 250 WORDS)

Activity-related rise in extracellular potassium concentration in the brain of 1-3-day-old chicks.

The concentration of K+ [( K+]) was measured in the cerebrospinal fluid (CSF) and in the extracellular fluid [( K+]e) in the medial forebrain of two-day-old chicks by means of K(+)-sensitive microelectrodes (K-ISM). The K-ISM potential in the CSF was compared with that in artificial CSF with 3, 4 or 6 mmol/l of [K+]. The [K+] found in CSF was 3.79 +/- 0.57 (n = 8), in the hyperstriatum accessorium (HA) 3.70 +/- 0.32 (n = 13) and in the neostriatum (N) 3.51 +/- 0.32 (n = 13; mmol/l; mean +/- SEM). Trains of local electrical stimuli (10-100 Hz, 30 sec) applied to the surface of the forebrain increased [K+]e in both HA and N by 11-13 mmol/l. Increases in [K+]e in the ectostriatum (E) of 5-6 mmol/l was found in response to electrical stimulation (30-100 Hz, 5-10 sec) of the contralateral optic nerve, and of about 2 mmol/l by applying pressure to the bulb. In chicks adapted to the dark, stimulation of the contralateral eye or both eyes with bright light flashes (1-2 Hz) or with continuous light resulted in an increase in [K+]e of 0.5-1.0 mmol/l. Smaller increases in [K+]e of 0.15-1.25 mmol/l were found in HV and N after the application of a small quantity (0.1 ml) of the chemical taste aversant methylanthranilate (MeA) or the electrical stimulation of the beak by two needle electrodes inserted into the palatum or into the tongue. After application of MeA the increase in [K+]e began 3-4 min after application and persisted for 20 min.(ABSTRACT TRUNCATED AT 250 WORDS)