Opposite membrane potential changes induced by glucose deprivation in striatal spiny neurons and in large aspiny interneurons

Abstract

We have studied the electrophysiological effects of glucose deprivation on morphologically identified striatal neurons recorded from a corticostriatal slice preparation. The large majority of the recorded cells were spiny neurons and responded to aglycemia with a slow membrane depolarization coupled with a reduction of the input resistance. In voltage-clamp experiments aglycemia caused an inward current. This current was associated with a conductance increase and reversed at -40 mV. The aglycemia-induced membrane depolarization was not affected by tetrodotoxin (TTX) or 6-cyano-7-nitroquinoxaline-2,3-dione plus aminophosphonovalerate, antagonists acting respectively on AMPA and NMDA glutamate receptors. Also, the intracellular injection of bis(2-aminophenoxy)ethane-N,N, N',N'-tetra-acetic acid, a calcium (Ca2+) chelator, and low Ca2+/high Mg2+-containing solutions failed to reduce this phenomenon. Conversely, it was reduced by lowering external sodium (Na+) concentration. A minority of the recorded cells had the morphological characteristics of large aspiny interneurons and the electrophysiological properties of "long-lasting afterhyperpolarization (LA) cells." These cells responded to aglycemia with a membrane hyperpolarization/outward current that was coupled with an increased conductance. This current was not altered by TTX, blockers of ATP-dependent potassium (K+) channels, and adenosine A1 receptor antagonists, whereas it was reduced by solutions containing low Ca2+/high Mg2+. This current reversed at -105 mV and was blocked by barium, suggesting the involvement of a K+ conductance. We suggest that the opposite membrane responses of striatal neuronal subtypes to glucose deprivation might account for their differential neuronal vulnerability to aglycemia and ischemia.

Fig. 1.

Morphological identification of striatal neurons. The figure represents two striatal neuronal subtypes: a biocytin-injected spiny striatal neuron (a) and a large LA interneuron filled with the dye fura-2 (b) (380 nm image, average of 256 frames) (for details, see Materials and Methods). Scale bar (shown in b): a, 70 μm;b, 30 μm.

Fig. 2.

Aglycemia depolarizes striatal spiny neurons and decreases their membrane input resistance. A, Thetop part represents selected chart records of the membrane potential changes caused by glucose deprivation. Theblack arrows indicate the onset of hypoglycemia (a), 15 min of aglycemia (b), the onset of the washout (c), and 10 min of washout (d). Interruptions of the traces between two open arrows represent different periods of recording: 13 min betweena and b, 13 min between band c, and 6 min between c andd. The bottom part shows the current trace monitored during the recording. The downward voltage deflections are induced by negative current steps. B, Single sweeps recorded at higher speed at the same times as those indicated inA. Current and voltage calibrations in Aapply also for B. In Bc, thedotted line indicates the original resting membrane potential (RMP = −85 mV). C, The tonic firing discharge (a) induced by a depolarizing pulse (b) in a striatal spiny cell (RMP= −86 mV).

Fig. 3.

Time course of the membrane depolarization induced by aglycemia in normal medium and in different experimental conditions. The graphs show the amplitude of the membrane depolarization induced by 25 min of glucose deprivation in control medium (filled circles), in 1 μmTTX (open circles), in 10 μm CNQX plus 50 μm APV (open rhombs), in low Na⁺ (38 mm, filled rhombs), in the presence of BAPTA-filled electrodes (open squares), in the presence of low calcium (0.5 mm)/high magnesium (10 mm) solutions (filled squares), and in the presence of the adenosine A1 receptor antagonist CPX (300 nm; filled triangles). Each data point represents the mean of at least four single observations.Asterisks indicate significant difference from control values (p < 0.01).

Fig. 4.

Aglycemia induces an inward current and increases the membrane conductance in spiny neurons. A, During a single-microelectrode voltage-clamp experiment, the voltage was monitored (see top part). The glucose deprivation induced an inward current. The black arrows indicate the onset of aglycemia (a), 15 min of aglycemia (b), the onset of the washout after 25 min of aglycemia (c), and 10 min of washout (d). Interruptions of the traces between two open arrowsrepresent different periods of recording: 14 min betweena and b, 7 min between band c, and 8 min between c andd. The downward deflections are induced by negative current steps. B, This part of the figure shows single sweeps recorded at higher speed at the same times as those indicated inA. Current and voltage calibrations in Aapply also for B. The holding potential during the experiment was −85 mV.

Fig. 5.

Characteristics of the inward current induced by aglycemia in spiny neurons. A, The graphshows the time course of the inward current induced by 25 min of glucose deprivation. B, The graph indicates the relative membrane conductance changes caused by 25 min of aglycemia.C, I–V relationship showing the extrapolated reversal potential (−40 mV, arrow) of the aglycemia-induced inward current. Long-lasting (1–3 sec) voltage steps were applied in both negative and positive directions before (filled circles) and during (open circles) aglycemia. The holding potential was −80 mV. Each data point represents the mean of at least four single observations.

Fig. 6.

Aglycemia hyperpolarizes LA striatal interneurons and decreases their membrane input resistance. A, Thetop part represents selected chart records of the membrane potential changes caused by glucose deprivation. Theblack arrows indicate the onset of aglycemia (a), 10 min of aglycemia (b), the onset of the washout (c), and 10 min of washout (d). Interruptions of the traces between two open arrows represent different periods of recording: 8 min betweena and b, 3 min between band c, and 7 min between c andd. The bottom part shows the current trace monitored during the recording. The downward voltage deflections are induced by negative current steps. B, Single sweeps recorded at higher speed at the same times as indicated inA. In Ac and Bc, thedotted line indicates the original resting membrane potential (−59 mV). C, A single action potential followed by a long-lasting hyperpolarization (a) is induced by a depolarizing current pulse (b) in an LA striatal interneuron. The dotted line represents the resting membrane potential of the neuron (−59 mV).

Fig. 7.

Aglycemia induces an outward current and increases the membrane conductance in LA striatal interneurons.A, During a single-microelectrode voltage-clamp experiment, aglycemia caused an outward current (toppart); during the experiment the voltage was monitored (seebottom part). The black arrows indicate the onset of aglycemia (a), the onset of the washout after 15 min of aglycemia (b), and 10 min after the washout (c). The interruption of the traces betweentwo open arrows represents a period of recording of 12 min. The downward deflections are induced by negative current steps.B, Single sweeps recorded at higher speed at the same times as indicated in A. The holding potential during the experiment was −60 mV.

Fig. 8.

The aglycemia-induced hyperpolarization/outward current in LA interneurons is mediated by a K⁺ conductance.A, Time course of the membrane changes induced by 25 min of glucose deprivation in different experimental conditions: control (n = 9; filled circles), 300 μm barium (n = 4; open circles), 1 mm tolbutamide (n = 3; open rhombs), 100 nm glipizide (n = 3; filled squares), 300 nm CPX (n = 4; open squares), and low calcium (0.5 mm)/high magnesium (10 mm) solutions (n = 4; filled rhombs). Asterisks indicate significant difference from control values (p < 0.01).B, In an LA interneuron, bath application of 30 μm adenosine induced a membrane hyperpolarization (a); this membrane hyperpolarization was fully blocked by 300 nm CPX, an adenosine A1 receptor antagonist (b); after the washout of this antagonist, the hyperpolarizing action of adenosine was restored (c).C, The reversal potential of the aglycemia-induced outward current is indicated by the arrow (−105 mV;n = 4). This value was calculated by measuring the steady-state currents generated by long-lasting (1–3 sec) voltage steps of progressively increasing amplitude from the holding potential (−60 mV; n = 4) before (filled circles) and during glucose deprivation (25 min, open circles).