H.D. Lux

Extracellular free calcium and potassium during paroxysmal activity in the cerebral cortex of the cat

SummaryExtracellular calcium and potassium activities (aCa and aK) as well as neuronal activity were simultaneously recorded with ion-sensitive electrodes in the somatosensory cortex of cats. Baseline aCa was 1.2–1.5 mM/1, baseline ak 2.7–3.2 mM/1. Transient decreases in aCa and simultaneous increases in aK were evoked by repetitive stimulation of the contralateral forepaw, the nucleus ventroposterolateralis thalami and the cortical surface. Considerable decreases in aCa (by up to 0.7 mM/1) were found during seizure activity. A fall in aCa preceded the onset of paroxysmal discharges and the rise in aK after injection of pentylene tetrazol. The decrease in aCa led also the rise in aK during cyclical spike driving in a penicillin focus. It is concluded that alterations of Ca++ dependent mechanisms participate in the generation of epileptic activity.

Relations between EEG phenomena and potentials of single cortical cells. I. Evoked responses after thalamic and epicortical stimulation

1. 1. The evoked EEG responses from the motor cortex of cats under Nembutal after electrical stimulation of nucleus VPL or VL of the thalamus, of midthalamic nuclei and of the cortex itself are compared with the cellular responses from the same area. The cellular responses were almost identical in different cells of a given experiment so that records from one cell can be taken as representative for the whole population of pyramidal cells from which the records were taken. The following correlations were found: 2. 2. VL stimulation: The primary surface positivity is accompanied at its peak by the primary EPSP which may lead to a discharge. If a secondary EPSP is recognizable, this is simultaneous with the surface negativity. The IPSP (present only during single stimuli or at medium frequencies at strong suprathreshold stimulation) begins during the surface positivity, and reaches its peak during the late surface positivity following the primary surface negativity. This positivity may continue as long as the IPSP lasts but is frequently surmounted by a late negativity (peak latency 30–50 msec). This corresponds to a third EPSP in the cellular record. The third as well as the secondary EPSP is frequently masked by the IPSP, especially during strong single stimuli. At medium stimulus frequencies both the late EPSPs as well as the surface negative responses are increased, whereas the IPSP and the secondary surface positivity decrease. At 15–20/sec stimulation the surface negativity becomes broader and the IPSP disappears. 3. 3. CM stimulation: The surface negative recruiting wave is accompanied by a summated cellular EPSP of the same time course. Waxing and waning is equally present in both records. Occasionally a positive notch on top of the recruiting wave is found which is accompanied by cellular discharges. The frequently observed slight surface positivity preceding the recruiting wave falls together with the first EPSPs; EEG arousal at high frequency stimulation is accompanied by corresponding cellular changes. 4. 4. Epicortical stimulation: The primary surface negativity (“superficial response”) is associated with a cellular excitation (direct excitation with spike discharge or EPSP) whereas the following surface positivity (deep response) comes together with the IPSP. This is especially clear in chronically isolated cortical slabs. IPSP and surface positivity have similar time courses. 5. 5. The cortical after-discharge after afferent or epicortical stimulation shows close relations between surface negative waves and cellular excitation. Some waves recorded with the surface electrode are not clearly related to cellular activity, which suggests that they are picked up from adjacent parts of the cortex. Further evidence is presented that the cortical after-discharge is due to thalamo-cortical activity. 6. 6. In the Discussion it is suggested that cortical evoked potentials are composed of different intracortical excitatory patterns: quick transients near the soma (with initial phase reversal in the surface record), superficially located, mostly slow dendritic potentials (without phase reversal), and synchronized afferent and efferent fiber activity.

Transient Changes in the Size of the Extracellular Space in the Sensorimotor Cortex of Cats in Relation to Stimulus-induced Changes in Potassium Concentration*

SummaryThe time course of local changes of the extracellular space (ES) was investigated by measuring concentration changes of repeatedly injected tetramethylammonium (TMA+) and choline (Ch+) ions for which cell membranes are largely impermeable. After stimulus-induced extracellular [K+] elevations the δ[TMA+] and δ[Ch+] signals recorded with nominally K+-selective liquid ion-exchanger microelectrodes increased by up to 100%, thus indicating a reduction of the ES down to one half of its initial size. The shrinkage was maximal at sites where the K+ release into the ES was also largest. At very superficial and deep layers, however, considerable increases in extracellular K+ concentration were not accompanied by significant reductions in the ES. These findings can be explained as a consequence of K+ movement through spatially extended cell structures. Calculations based on a model combining the spatial buffer mechanism of Kuffler and Nicholls (1966) to osmolarity changes caused by selective K+ transport through primarily K+ permeable membranes support this concept.Following stimulation additional iontophoretically induced [K+]o rises were reduced in amplitude by up to 35%, even at sites where maximal decreases of the ES were observed. This emphasizes the importance of active uptake for K+ clearance out of the ES.

The equilibration time course of (K + ) 0 in cat cortex.

SummarySteady state and transient values of intracortical potassium were measured with K+ sensitive microelectrodes. Resting intracortical K+ activity is low and resembles that of cerebrospinal fluid. Elevation of intracortical K+ was brought about by electrophoretic injection of K+ by a constant current source from a KCl containing micropipette at fixed distances from the recording electrode. The intracortical K+ responses to electrophoretic K+ injection were compared with those in a medium of 150 mM/l NaCl plus 3 mM/l KCl. The dependence of intracortical K+ steady state levels on electrophoretic currents is nearly linear, but the K+ response in the cortex was about six times higher than in saline. Half times (T1/2) of the rising and falling phases of K+ during current steps were found to be prolonged by the same degree in the cortex. The distribution of [K+]0 appears to be dominated by free diffusion with an apparent diffusion coefficient of 1/6 that in the medium. Primarily diffusional redistribution may also apply to K+ which is released by direct cortical stimulation. K+ released by brief stimulation distributes faster than K+ during and after prolonged continuous stimulation with average T1/2 of 1.2 and 3.0 sec respectively in accordance with diffusion from instantaneous and continuous point sources. For small [K+]0 changes, deviations from diffusional kinetics were found to be about one-fifth of absolute [K+]0 values and became predominant at times longer than 10 T1/2. They can be ascribed to K+ uptake mechanisms. DC recorded cortical surface potentials reveal close relations to the slopes of intracortical potassium activity.

Effects of dopamine and noradrenaline on Ca channels of cultured sensory and sympathetic neurons of chick

The effects of noradrenaline and dopamine on voltage-dependent Ca currents were investigated in cultured dorsal root and sympathetic ganglion neurons from chick embryos. At concentrations of 1 to 10 μM, bath application of the neurotransmitters caused a general depression of inward Ca currents. Above −20 mV the decrease of the current amplitude was reversible and accompanied by a 2–10-fold prolongation of the activation time course. Below −20 mV, where a low voltage-activated Ca component is turned on, the size of the currents was reduced by 40% with little effect on the time course. Despite extensive wash-out, little sign of reversibility was observed in this case.Single-channel current recording in outside-out membrane patches revealed that a low membrane potentials dopamine and noradrenaline reversibly reduced single Ca-channel activity. This finding supports the view that in sensory and sympathetic neurons, both neurotransmitters affect the membrane conductance by modulating Ca permeability and not by activating catecholamine-specific channels able to carry transient outward currents. The probability of Ca channel opening is strongly reduced by addition of 10 μM of either catecholamine to the bath. The possible involvement of a voltage-dependent block of Ca channels by the neurotransmitters is discussed.

Relations between EEG phenomena and potentials of single cortical cells. II. Spontaneous and convulsoid activity.

Abstract 1. 1. The relations between spontaneous EEG waves, convulsoid potentials (induced by i.v. Metrazol injection or epicortical stimulation) and intracellularly recorded activity were analyzed in the motor cortex of cats under Nembutal. 2. 2. During regular surface negative spindle waves a close positive correlation exists between cellular depolarization and surface negativity. The cellular depolarizations are composed of several EPSPs which can be easily distinguished in some cells. Sometimes they begin to summate during a slight surface positivity preceding the negative wave. The summated post-synaptic depolarization is preceded by a period of synaptic silence, indicating an extracellular, probably extracortical, trigger mechanism. IPSPs are seen only if much spike activity is present. 3. 3. Another type of spindle wave (sometimes seen in isolation) is characterized by a relatively short and weak surface negative wave followed by a longer positive potential. The cellular activity shows more or less pronounced depolarization and spike activity during or just preceding the negative wavelet and an IPSP during the surface positive potential. The IPSP may begin during the peak or falling phase of the surface negative potential. The presence of active inhibition in these cases has been proved by intracellular stimulation. 4. 4. If biphasic positive-negative spindle waves are present their relation to the cellular activity depends upon the potential gradient of the wave. If the positive-negative gradient is steep, cellular activity with a relatively high discharge rate is located on the positive phase; if the gradient is relatively small the slow summation of EPSPs reaches its peak during the negative wave (see Fig. 9). 5. 5. At the beginning of Metrazol-induced seizures slow surface negative waves occur simultaneously with cellular depolarization. During the peak of the seizure, when mainly large biphasic positive-negative potentials are seen in the EEG, massive cellular depolarization, leading quickly to high frequency grouped spike discharges, occurs with the first surface positive phase. Cellular repolarization coincides with the positive-negative transition of the surface potential. The biphasic EEG potential is frequently followed by a longer positive potential which corresponds to a cellular IPSP. At the end of the seizure isolated IPSPs are frequently seen. 6. 6. Strong high frequency epicortical stimulation leads to a long lasting depolarization of cortical cells (down to 20–30 mV) with inactivation of the spike generator. After the end of the stimulation the membrane potential shows regular oscillations, the spike generator recovers and the cell polarizes slowly. Finally, oscillating depolarizations are interrupted by long polarizations. Some cells do not show a long lasting depolarization after the stimulus but are subsequently synaptically depolarized during the depolarizing oscillations. The depolarizations occur simultaneously with large surface positive potentials of the cortical after-discharge. 7. 7. Possible mechanisms relating EEG waves to cellular potentials are discussed within the framework of transcortical and soma-apical dendrite potential distributions during different phases of physiological and pathological cortical activity.

Stimulus-induced changes in extracellular Na + and Cl − concentration in relation to changes in the size of the extracellular space

SummaryExtracellular Na+- and Cl−-concentrations ([Na+]o, [Cl−]o) were recorded with ion-selective microelectrodes during repetitive stimulation and stimulus-induced self-sustained neuronal afterdischarges (SAD) in the sensorimotor cortex of cats. In all cortical layers [Na+]o initially decreased by 4–7 mM. In depths of more than 600 μm below the cortical surface such decreases usually turned into increases of 2–6 mM during the course of the SADs, whereas in superficial layers [Na+]o never rose above its resting level. [Cl−]o always showed an increase in the course of the SADs often preceded by an initial small decrease. The average increase at a depth of 1,000 μm was about 7 mM. [Cl−]o reached peak values at about the end of the ictal period, whereas [Na+]o reached its maximum shortly after the end of the SAD, at times when [K+]o was still elevated above the baseline concentration.These data indicate that the extracellular osmolarity can increase during SAD by up to 30 mM. Such an increase in osmolarity can be explained by an increase in the number of intracellular particles, caused by cleavage of larger molecules during enhanced metabolism. This could lead to cell-swelling due to passive water influx from the extracellular space (ES). However, the resulting reduction of the size of the ES is calculated to be less than 10% for an increase in intracellular osmolarity by 30 mOsm. This value is too small as compared to previously measured ES-reductions under similar conditions (i.e., 30% reduction at 1,000 μm; Dietzel et al. 1980). Reductions of the size of the ES that accompany the observed changes in the ionic environment, are quantitatively explained on the basis of the extended glial buffering mechanism described in the preceding paper.

Undershoots following stimulus-induced rises of extracellular potassium concentration in cerebral cortex of cat.

Extracellular potassium activity (ak) was recorded with potassium-sensitive electrodes in the sensorimotor cortex of cats. Resting activity was 2.8--3.4 mEquiv/l. Electric stimulation of the cortical surface and the nucleus ventroposterolateralis of the thalamus brought about an increase in aK followed by an undershoot and return to normal value. The lowest observed value of aK was 2.1 mEquiv./l. Size and duration (range 0.5--4 min) of the undershoots of aK increased with increasing peak amplitudes of the preceding rise in aK. Following the rise in aK, a period of reduced neuronal activity was observed which usually shorter lasting than the decrease in extracellular aK. An undershoot of aK and a concomitant reduction of neuronal discharge frequency can also occur in immediate response to antidromic stimulation of the pyramidal tract. To compare the K+ redistribution at normal and reduced levels of aK electrophoretic K+ signals were produced with constant current pulses from a proximate KCl-filled capillary. Both amplitudes and half times of decay of these K+ signals were found to decrease during the phase of poststimulatory undershoot in aK (19 and 23% respectively). It is suggested that an activated reuptake of potassium contributes to the decrease in extracellular aK in addition to inhibitory processes.

γ-Aminobutyric acid-induced depression of calcium currents of chick sensory neurons

The effects of gamma-aminobutyric acid (GABA) on calcium currents were investigated in avian dorsal root ganglion (DRG) cells. GABA was applied to the vicinity of the cells by ejection pipettes using constant-pressure pulses. GABA concentrations between 5 and 100 mu M reduced and slowed the calcium current in a dose-dependent manner. A contribution of K and Cl outward currents to the reduction of the inward current was minimized by using identical caesium chloride concentrations on both sides of the membrane. The onset of the effect was rapid and 80% of the effect was observed within 1 s. The attenuation of the Ca slope conductance by GABA was found to be independent of the membrane potential between -50 and +50 mV.

Excitation and axonal flow: Autoradiographic study on motoneurons intracellularly injected with a 3H-amino acid

SummaryTritiated glycine has been injected intracellularly into cat motoneurons by means of cross barrel iontophoresis through microelectrodes. The amino acid is incorporated into proteins in the cell body and the synthesized radioactive proteins have been localized by autoradiography. The single-cell-injection technique has several advantages over the usual intraperitoneal, intravenous or intrathecal injection, e.g. distribution of the radiochemical is strictly confined to the injected neuron; the amount of injected substance can be estimated; it is easy to investigate simultaneously electrophysiological and morphological characteristics of a defined neuron. Since the injected cell body releases radioactive proteins into the axon, the transport of these substances can be studied under various conditions.In this study the influence of antidromic stimulation has been studied. Stimulated motoneurons demonstrate higher radioactivity in their cell bodies than unstimulated ones, reflecting an increased protein synthesis. The differences are even more pronounced in the axons of stimulated cells when compared with unstimulated ones. A significant, higher amount of radioactive material — up to 100% increase as demonstrated by silver grain counting — can be seen in the axons of stimulated neurons. Although considerable differences seem to exist for the quantity of the exported proteins no significant differences have been detected for the velocity of transportation. In axons of stimulated and unstimulated neurons proteins advance toward the periphery at the same rate.