Neuroscience Letters, 133 (1991) 93-96
93
© 1991 Elsevier Scientific Publishers Ireland Ltd. All rights reserved 0304-3940/91/$ 03.50 NSL 08202
5-Hydroxytryptamine hyperpolarizes CA3 hippocampal pyramidal cells through an increase in potassium conductance Sheryl G. Beck and Kue C. Choi Department of Pharmacology, Loyola University Chicago, Stritch School of Medicine, Maywood, IL (U.S.A.) (Received 7 July 1991; Revised version received 19 August 1991; Accepted 20 August 1991)
Key words: Serotonin; Hippocampal slice; Potassium; Hyperpolarization; 5-Hydroxytryptamine; Intracellular; CA3 The firing rate of hippocampal pyramidal cells recorded from the CA3 subfield is inhibited by 5-hydroxytryptamine (5-HT, serotonin) or by electrical stimulation of the ascending serotonergic fibers from the raphe. The mechanism of action of this inhibitory effect produced by 5-HT has not been determined. Intracellular recording techniques in the hippocampal slice preparation were used to measure the effect of 5-HT perfusion on membrane properties of CA3 pyramidal cells. In 15 out of 16 cells tested, 5-HT elicited a pronounced hyperpolarization concomitant with a decrease in membrane resistance. The hyperpolarization was not altered with either potassium chloride or potassium methylsulphate electrodes; the hyperpolarization by 5-HT was not present when electrodes were filled with cesium chloride. The reversal potential of the 5-HT mediated response was determined to be - 105.5 mV in 3 mM KCI buffer using single electrode voltage clamp techniques. Based on these results we conclude that the mechanism of action of the 5-HT inhibition of CA3 hippocampal pyramidal cell excitability is due to an increase in potassium conductance.
All subfields of the hippocampus receive extensive innervation from 5-hydroxytryptamine (5-HT, serotonin) cell bodies in the raphe [2, 8, 9]. Serotonin binding sites, visualized by autoradiographic techniques, are found in high density in area CA1 and the dentate gyrus, but in lower density in area CA3 [11, 16]. Previous studies, using extracellular single unit recording techniques, reported that 5-HT elicits a reduction in CA1 and CA3 cell firing rate [7, 10, 12, 14]. This effect of 5-HT is mimicked by stimulating the ascending serotonergic fibers [5, 13]. In area CAI of the hippocampus, 5-HT elicits a hyperpolarization due to an increase in potassium conductance. This hyperpolarization by 5-HT has been extensively studied and the receptor mediating this response has been identified as the 5-HTIA receptor [1, 3, 6]. In contrast, the mechanism of action of 5-HT in area CA3 has not been rigorously studied. One paper reported that 5-HT hyperpolarized CA3 hippocampal pyramidal cells with little or no change in membrane resistance [15]. Since a reversal potential was not apparent, the author postulated that 5-HT hyperpolarized CA3 pyramidal cells through a mechanism other than an increase in Correspondence: S.G. Beck, Department of Pharmacology, Loyola University Chicago, Stritch School of Medicine, 2160 South First Avenue, Maywood, IL 60153, U.S.A. Fax: (1) (708) 216-4118.
potassium conductance. We report here that 5-HT elicits an hyperpolarization of CA3 hippocampal pyramidal cells through an increase in potassium conductance. Methods were as previously described [4]. Briefly, male Sprague--Dawley rats (75-200 g) were anesthetized with ether and decapitated. The brain was rapidly removed and rinsed in ice cold artificial cerebrospinal fluid (ACSF). The composition of the ACSF was (in mM): NaCI 124, KC1 3, NaH2PO4 1.25, MgSO4 2, CaC1z 2.5, dextrose 10 NaHCO 3 28 and bubbled with 95% 02/ 5% CO2 to maintain the pH at 7.4. The right dorsal hippocampus was dissected free and 500-600/zm sections were cut on a vibratome starting from the septal/dorsal tip. The slices were placed in a holding chamber containing ACSF bubbled with 95% 02/5% CO2 to maintain pH at 7.4. After approximately one hour a slice was transfered to a recording chamber where it was sandwiched between two nylon nets, submerged in ACSF (32 + 1°C) and continuously perfused at a flow rate of 2-3 ml/min. 5-HT was applied by bath perfusion in known concentrations. Standard intracellular recording techniques were used. Electrodes were pulled from borosilicate capillary tubing (1.2 mm o.d., 0.69 mm i.d.) to obtain resistances of 40140 m£2 (2 M KCI, 2 M KCH3SO4 or 2 M CsC1). Hippocampal pyramidal cells in area CA3 were impaled by briefly increasing the capacity compensation or by inject-
94 ing a large depolarizing current. Electrical signals were amplified, recorded on a chart recorder, digitized and stored for later data analysis using an IBM compatible computer system and Axoclamp software (Axon Instruments). Statistical analysis included analysis of variance and Students t-test; a P < 0.05 was considered significant. Data for concentration-response curves were fit to a hyperbolic function where E = Em~x/(I+(ECso/[5HT])N): E is the response elicited by the 5-HT concentration [5-HT], ECs0 is the concentration of 5-HT that produces a response 50% of maximum, Em~, and N is equal to the slope. Using this formula estimates for Em~x, ECso and N were obtained using RS/1 software (Bolt Beranek and Neuman). The reversal potential of the 5-HT induced current was measured using single electrode voltage clamp techniques. The cells were clamped at their resting membrane potential. Electrodes were filled with 2 M KC1 and the fluid level over the slice adjusted to less than 200 pm. For some cells, the tips of the electrodes were coated with M Coat D (Measurement Group Inc., Raleigh, NC) to reduce capacitance. The headstage was continuously monitored and the switching frequency set at 4-8 kHz to allow for full discharge of the electrode between cycles. Data for I - V plots was obtained by hyperpolarizing the membrane potential in 10 mV increments and measuring the amount of current elicited. A maximal concentration of 5-HT (usually 100/~M) was added to the ACSF while holding the cell at its resting membrane potential and the amount of outward current measured.
5-HT
3
6
10
Once steady-state current change was reached, data for an I-V plot in the presence of 5-HT was obtained. The data for a cell was not used if the membrane potential deviated more than 2 mV from the clamped potential during 5-HT perfusion. Chemicals for the ACSF were purchased from Fisher Scientific and 5-hydroxytryptamine hydrochloride was purchased from Sigma. Data were collected from 23 cells in area CA3. The average resting membrane potential was - 6 5 . 3 _+4.9 mV (mean +S.D.), membrane resistance was 61.9 +11.8 MI2, and action potential height was 89.7 + 4.1 mV. In 15 out of 16 cells, perfusion of the slice with 5-HT elicited a hyperpolarization greater than 6 mV (range 6-24 mV) that was reversible and concentration-dependent. Upon repeated administration of the same 5-HT concentration, the magnitude of the response was the same, i.e., there was no tachyphylaxis. Fig. 1 contains a chart recording of a CA3 cell depicting the concentration dependent response to 5-HT perfusion. Estimates of Em~x, ECs0 and slope were obtained for each cell. The mean Emax was a hyperpolarization of 14.9 +5.1 mV, ECso was 10 +5/~M, and slope was 1.8 +0.6 (n = 15). Fig. 2 contains the summary concentration-response curve to 5-HT for all of the CA3 cells. The Emax for CA3 cells appeared to be larger than that reported for 5-HT in area CA 1 of the hippocampus, which was approximately 10-12 mV [1, 4]. Also, 5-HT appeared to be less potent in area CA3 since the ECs0 was shifted about a half log unit to the right as compared to CA1 [1, 4].
30
(uM)
;lmlllr;ml]il1lm!,i ,,
,,
1
O0
i
]lOmV 2min
Fig. 1. Continuous chart recordingof a CA3 hippocampal pyramidal cell response to perfusion by increasingconcentrationsof 5-HT. The resting membrane potential for this cell was -61 mV. The length of the line at the top of the figure reflectsthe duration of 5-HT perfusion for the concentration indicated. The downward deflectionsrepresent the change in membrane potential in response to a -300 pA, 300 ms current injection through the recordingelectrode.
95 20
A
A
500 •
(9)
I 250 •
N
1,4
Control 5HT
o
/ /
¢I
10
I-I 1.4 m O
0
@h
(
-250
-
-500
•
-750
•
Potential
9~I
(15V.L / 0
. . . . . (12) -./ 10 -7
10 -6
J l• ......,
........ ,
i0 -5
[ 5-HT]
10 -4
........ , 10 -3
-I000.
(M)
Fig. 2. Summary graph o f the concentration-dependent nature o f the 5-HT hyperpolarization. The number in parentheses above each data point represents the number o f cells tested at each concentration. The bars are __+the standard error of the mean.
Segal [15] previously reported that there was little or no change in membrane resistance in response to 5-HT perfusion, especially as compared to the response in CA1, and that there was no intersection in the graph of the control and 5-HT I - V plots using bridge balance recording conditions. Based on this data, Segal concluded that the 5-HT mediated hyperpolarization was not due to an increase in potassium conductance. In the experiments reported here, there was an overall 35% reduction in membrane resistance, from 61.9 + 11.8 M[2 to 40.1 +20 Ml2, during 5-HT perfusion (P = 0.0109, Students two-way paired t-test, n = 14). A series of experiments were conducted to test whether the hyperpolarization elicited by 5-HT in area CA3 was due to an increase in potassium conductance. 5-HT elicited a hyperpolarization with either KC1 (n = 11) or KCH3SO + (n = 4) electrodes, ruling out the possibility that the hyperpolarization was due to a change in chloride conductance. When recording electrodes were filled with 2 M CsC1, 5-HT (100/zM) elicited no hyperpolarization (n = 3), implicating a change in potassium conductance. The hyperpolarization was not altered in the presence of the sodium channel blocker tetrodotoxin at 1/zM (n = 2). Single electrode voltage clamp techniques were used to measure the reversal potential for the 5-HT elicited response. Fig. 3A contains the 1-V plots constructed from data collected in the absence and presence of 100/IM 5HT for a cell recorded under voltage clamp. Fig. 3B contains a graph of the 5-HT induced current, obtained by subtracting the control values from the values obtained during 5-HT perfusion. The reversal potential for this
B 200
-
i00
'
v
¢ bl 1.4 m O
i
-120
•
/
i
y
i
-i00
-90
-80
-70
Potential (mY)
-i00
Fig. 3. A: I - V plots taken from data collected in the absence (control) and presence o f 100/~M 5-HT under voltage clamp. B: the amount o f current elicited by 5-HT perfusion at different membrane potentials. This I - V plot was obtained by subtracting the control values from the 5-HT values shown in A. The reversal potential was - 107 mV.
cell was - 1 0 7 mV; for 4 cells with 3 mM KCI in the ACSF the reversal potential was - 106.7 + 1.5 mV. This value is similar to the reversal potential reported for 5HT in area CA1 in the presence of 2.5 or 3 mM extracellular potassium [1, 6]. From these results we conclude that there is a pronounced hyperpolarization to 5-HT in area CA3 of the hippocampus which is due to an increase in potassium conductance. This hyperpolarization probably underlies the decrease in cell firing measured using extracellular single unit recording techniques [7, 10]. Further studies are currently being conducted to characterize the 5-HT receptor mediating the hyperpolarization in area CA3. Also under investigation are the differences in the nature of the hyperpolarizing response recorded from cells in areas CA1 and CA3.
96 T h i s w o r k was s u p p o r t e d in p a r t by g r a n t s f r o m the P o t t s Estate, L o y o l a U n i v e r s i t y C h i c a g o S t r i t c h S c h o o l of Medicine, PHS
Grant
Scientist D e v e l o p m e n t NIMH
NS-28512
Award,
and a Research
KO2-MH-00880
from
to S G B .
1 Andrade, R. and Nicoll, R.A., Pharmacologically distinct actions of serotonin on single pyramidal neurones of the rat hippocampus recorded in vitro, J. Physiol., 394 (1987) 99-124. 2 Azmitia, E.C. and Segal, M., An autoradiographic analysis of the differential ascending projections of the dorsal and median raphe nuclei in the rat, J. Comp. Neurol., 179 (1978) 641-668. 3 Beck, S.G., 5-Carboxyamidotryptamine mimics only the 5-HT elicited hyperpolarization of hippocampal pyramidal cells via 5-HTtA receptor, Neurosci. Lett., 99 (1989) 101-106. 4 Beck, S.G. and Halloran, P.H., lmipramine alters fl-adrenergic, but not serotonergic, mediated responses in rat hippocampal pyramidal cells, Brain Res., 504 (1989) 72-81. 5 Chaput, Y., Blier, P. and deMontigny, C., In vivo electrophysiological evidence for the regulatory role ofautoreceptors on serotonergic terminals, J. Neurosci., 6 (1986) 2796- 2801. 6 Colino, A. and Halliwell, J.V., Differential modulation of three separate K-conductances in hippocampal CAI neurons by serotonin, Nature, 328 (1987) 73 77. 7 deMontigny, C. and Aghajanian, G.K., Tricyclic antidepressants: long-term treatment increases responsivity of rat forebrain neurons to serotonin, Science, 202 (1978) 1303 1306.
8 Molliver, M.E., Serotonergic neuronal systems: what their anatomic organization tells us about function, J. Clin. Psychopharmacol., 7 (1987) 3S-23S. 90leskevich, S. and Descarries, L., Quantified distribution of the serotonin innervation in adult rat hippocampus, Neuroscience, 34 (1990) 19--33. 10 Otmakhov, N.A. and Bragin, A.G., Effects of norepinephrine and serotonin upon spontaneous activity and responses to mossy fiber stimulation of CA3 neurons in hippocampal slices, Brain Res., 253 (1982) 173-183. 11 Pazos, A. and Palacios, J.M., Quantitative autoradiographic mapping of serotonin receptors in the rat brain. I. Serotonin-1 receptors, Brain Res., 346 (1985) 205 230. 12 Penington, N.J. and Reiffenstein, R.J., Lack of effect of antagonists on serotonin-induced inhibition in rat hippocampus, Can. J. Physiol. Pharmacol., 64 (1986) 1413-1418. 13 Segal, M., Physiological and pharmacological evidence for a serotonergic projection to the hippocampus, Brain Res., 94 (1975) 115131. 14 Segal, M., 5-HT antagonists in rat hippocampus, Brain Res., 103 (1976) 161--166. 15 Segal, M., Regional differences in neuronal responses to 5-HT: intracellular studies in hippocampal slices, J. Physiol., 77 (1981) 373 -375. 16 Wilner, S.A., deMontigny, C., Desroches, J., Desjardins, P. and Suranyi-Cadotte, B.E., Autoradiographic quantification of serotonin~A receptors in rat brain following antidepressant drug treatment, Synapse, 4 (1989) 347 352.
93
© 1991 Elsevier Scientific Publishers Ireland Ltd. All rights reserved 0304-3940/91/$ 03.50 NSL 08202
5-Hydroxytryptamine hyperpolarizes CA3 hippocampal pyramidal cells through an increase in potassium conductance Sheryl G. Beck and Kue C. Choi Department of Pharmacology, Loyola University Chicago, Stritch School of Medicine, Maywood, IL (U.S.A.) (Received 7 July 1991; Revised version received 19 August 1991; Accepted 20 August 1991)
Key words: Serotonin; Hippocampal slice; Potassium; Hyperpolarization; 5-Hydroxytryptamine; Intracellular; CA3 The firing rate of hippocampal pyramidal cells recorded from the CA3 subfield is inhibited by 5-hydroxytryptamine (5-HT, serotonin) or by electrical stimulation of the ascending serotonergic fibers from the raphe. The mechanism of action of this inhibitory effect produced by 5-HT has not been determined. Intracellular recording techniques in the hippocampal slice preparation were used to measure the effect of 5-HT perfusion on membrane properties of CA3 pyramidal cells. In 15 out of 16 cells tested, 5-HT elicited a pronounced hyperpolarization concomitant with a decrease in membrane resistance. The hyperpolarization was not altered with either potassium chloride or potassium methylsulphate electrodes; the hyperpolarization by 5-HT was not present when electrodes were filled with cesium chloride. The reversal potential of the 5-HT mediated response was determined to be - 105.5 mV in 3 mM KCI buffer using single electrode voltage clamp techniques. Based on these results we conclude that the mechanism of action of the 5-HT inhibition of CA3 hippocampal pyramidal cell excitability is due to an increase in potassium conductance.
All subfields of the hippocampus receive extensive innervation from 5-hydroxytryptamine (5-HT, serotonin) cell bodies in the raphe [2, 8, 9]. Serotonin binding sites, visualized by autoradiographic techniques, are found in high density in area CA1 and the dentate gyrus, but in lower density in area CA3 [11, 16]. Previous studies, using extracellular single unit recording techniques, reported that 5-HT elicits a reduction in CA1 and CA3 cell firing rate [7, 10, 12, 14]. This effect of 5-HT is mimicked by stimulating the ascending serotonergic fibers [5, 13]. In area CAI of the hippocampus, 5-HT elicits a hyperpolarization due to an increase in potassium conductance. This hyperpolarization by 5-HT has been extensively studied and the receptor mediating this response has been identified as the 5-HTIA receptor [1, 3, 6]. In contrast, the mechanism of action of 5-HT in area CA3 has not been rigorously studied. One paper reported that 5-HT hyperpolarized CA3 hippocampal pyramidal cells with little or no change in membrane resistance [15]. Since a reversal potential was not apparent, the author postulated that 5-HT hyperpolarized CA3 pyramidal cells through a mechanism other than an increase in Correspondence: S.G. Beck, Department of Pharmacology, Loyola University Chicago, Stritch School of Medicine, 2160 South First Avenue, Maywood, IL 60153, U.S.A. Fax: (1) (708) 216-4118.
potassium conductance. We report here that 5-HT elicits an hyperpolarization of CA3 hippocampal pyramidal cells through an increase in potassium conductance. Methods were as previously described [4]. Briefly, male Sprague--Dawley rats (75-200 g) were anesthetized with ether and decapitated. The brain was rapidly removed and rinsed in ice cold artificial cerebrospinal fluid (ACSF). The composition of the ACSF was (in mM): NaCI 124, KC1 3, NaH2PO4 1.25, MgSO4 2, CaC1z 2.5, dextrose 10 NaHCO 3 28 and bubbled with 95% 02/ 5% CO2 to maintain the pH at 7.4. The right dorsal hippocampus was dissected free and 500-600/zm sections were cut on a vibratome starting from the septal/dorsal tip. The slices were placed in a holding chamber containing ACSF bubbled with 95% 02/5% CO2 to maintain pH at 7.4. After approximately one hour a slice was transfered to a recording chamber where it was sandwiched between two nylon nets, submerged in ACSF (32 + 1°C) and continuously perfused at a flow rate of 2-3 ml/min. 5-HT was applied by bath perfusion in known concentrations. Standard intracellular recording techniques were used. Electrodes were pulled from borosilicate capillary tubing (1.2 mm o.d., 0.69 mm i.d.) to obtain resistances of 40140 m£2 (2 M KCI, 2 M KCH3SO4 or 2 M CsC1). Hippocampal pyramidal cells in area CA3 were impaled by briefly increasing the capacity compensation or by inject-
94 ing a large depolarizing current. Electrical signals were amplified, recorded on a chart recorder, digitized and stored for later data analysis using an IBM compatible computer system and Axoclamp software (Axon Instruments). Statistical analysis included analysis of variance and Students t-test; a P < 0.05 was considered significant. Data for concentration-response curves were fit to a hyperbolic function where E = Em~x/(I+(ECso/[5HT])N): E is the response elicited by the 5-HT concentration [5-HT], ECs0 is the concentration of 5-HT that produces a response 50% of maximum, Em~, and N is equal to the slope. Using this formula estimates for Em~x, ECso and N were obtained using RS/1 software (Bolt Beranek and Neuman). The reversal potential of the 5-HT induced current was measured using single electrode voltage clamp techniques. The cells were clamped at their resting membrane potential. Electrodes were filled with 2 M KC1 and the fluid level over the slice adjusted to less than 200 pm. For some cells, the tips of the electrodes were coated with M Coat D (Measurement Group Inc., Raleigh, NC) to reduce capacitance. The headstage was continuously monitored and the switching frequency set at 4-8 kHz to allow for full discharge of the electrode between cycles. Data for I - V plots was obtained by hyperpolarizing the membrane potential in 10 mV increments and measuring the amount of current elicited. A maximal concentration of 5-HT (usually 100/~M) was added to the ACSF while holding the cell at its resting membrane potential and the amount of outward current measured.
5-HT
3
6
10
Once steady-state current change was reached, data for an I-V plot in the presence of 5-HT was obtained. The data for a cell was not used if the membrane potential deviated more than 2 mV from the clamped potential during 5-HT perfusion. Chemicals for the ACSF were purchased from Fisher Scientific and 5-hydroxytryptamine hydrochloride was purchased from Sigma. Data were collected from 23 cells in area CA3. The average resting membrane potential was - 6 5 . 3 _+4.9 mV (mean +S.D.), membrane resistance was 61.9 +11.8 MI2, and action potential height was 89.7 + 4.1 mV. In 15 out of 16 cells, perfusion of the slice with 5-HT elicited a hyperpolarization greater than 6 mV (range 6-24 mV) that was reversible and concentration-dependent. Upon repeated administration of the same 5-HT concentration, the magnitude of the response was the same, i.e., there was no tachyphylaxis. Fig. 1 contains a chart recording of a CA3 cell depicting the concentration dependent response to 5-HT perfusion. Estimates of Em~x, ECs0 and slope were obtained for each cell. The mean Emax was a hyperpolarization of 14.9 +5.1 mV, ECso was 10 +5/~M, and slope was 1.8 +0.6 (n = 15). Fig. 2 contains the summary concentration-response curve to 5-HT for all of the CA3 cells. The Emax for CA3 cells appeared to be larger than that reported for 5-HT in area CA 1 of the hippocampus, which was approximately 10-12 mV [1, 4]. Also, 5-HT appeared to be less potent in area CA3 since the ECs0 was shifted about a half log unit to the right as compared to CA1 [1, 4].
30
(uM)
;lmlllr;ml]il1lm!,i ,,
,,
1
O0
i
]lOmV 2min
Fig. 1. Continuous chart recordingof a CA3 hippocampal pyramidal cell response to perfusion by increasingconcentrationsof 5-HT. The resting membrane potential for this cell was -61 mV. The length of the line at the top of the figure reflectsthe duration of 5-HT perfusion for the concentration indicated. The downward deflectionsrepresent the change in membrane potential in response to a -300 pA, 300 ms current injection through the recordingelectrode.
95 20
A
A
500 •
(9)
I 250 •
N
1,4
Control 5HT
o
/ /
¢I
10
I-I 1.4 m O
0
@h
(
-250
-
-500
•
-750
•
Potential
9~I
(15V.L / 0
. . . . . (12) -./ 10 -7
10 -6
J l• ......,
........ ,
i0 -5
[ 5-HT]
10 -4
........ , 10 -3
-I000.
(M)
Fig. 2. Summary graph o f the concentration-dependent nature o f the 5-HT hyperpolarization. The number in parentheses above each data point represents the number o f cells tested at each concentration. The bars are __+the standard error of the mean.
Segal [15] previously reported that there was little or no change in membrane resistance in response to 5-HT perfusion, especially as compared to the response in CA1, and that there was no intersection in the graph of the control and 5-HT I - V plots using bridge balance recording conditions. Based on this data, Segal concluded that the 5-HT mediated hyperpolarization was not due to an increase in potassium conductance. In the experiments reported here, there was an overall 35% reduction in membrane resistance, from 61.9 + 11.8 M[2 to 40.1 +20 Ml2, during 5-HT perfusion (P = 0.0109, Students two-way paired t-test, n = 14). A series of experiments were conducted to test whether the hyperpolarization elicited by 5-HT in area CA3 was due to an increase in potassium conductance. 5-HT elicited a hyperpolarization with either KC1 (n = 11) or KCH3SO + (n = 4) electrodes, ruling out the possibility that the hyperpolarization was due to a change in chloride conductance. When recording electrodes were filled with 2 M CsC1, 5-HT (100/zM) elicited no hyperpolarization (n = 3), implicating a change in potassium conductance. The hyperpolarization was not altered in the presence of the sodium channel blocker tetrodotoxin at 1/zM (n = 2). Single electrode voltage clamp techniques were used to measure the reversal potential for the 5-HT elicited response. Fig. 3A contains the 1-V plots constructed from data collected in the absence and presence of 100/IM 5HT for a cell recorded under voltage clamp. Fig. 3B contains a graph of the 5-HT induced current, obtained by subtracting the control values from the values obtained during 5-HT perfusion. The reversal potential for this
B 200
-
i00
'
v
¢ bl 1.4 m O
i
-120
•
/
i
y
i
-i00
-90
-80
-70
Potential (mY)
-i00
Fig. 3. A: I - V plots taken from data collected in the absence (control) and presence o f 100/~M 5-HT under voltage clamp. B: the amount o f current elicited by 5-HT perfusion at different membrane potentials. This I - V plot was obtained by subtracting the control values from the 5-HT values shown in A. The reversal potential was - 107 mV.
cell was - 1 0 7 mV; for 4 cells with 3 mM KCI in the ACSF the reversal potential was - 106.7 + 1.5 mV. This value is similar to the reversal potential reported for 5HT in area CA1 in the presence of 2.5 or 3 mM extracellular potassium [1, 6]. From these results we conclude that there is a pronounced hyperpolarization to 5-HT in area CA3 of the hippocampus which is due to an increase in potassium conductance. This hyperpolarization probably underlies the decrease in cell firing measured using extracellular single unit recording techniques [7, 10]. Further studies are currently being conducted to characterize the 5-HT receptor mediating the hyperpolarization in area CA3. Also under investigation are the differences in the nature of the hyperpolarizing response recorded from cells in areas CA1 and CA3.
96 T h i s w o r k was s u p p o r t e d in p a r t by g r a n t s f r o m the P o t t s Estate, L o y o l a U n i v e r s i t y C h i c a g o S t r i t c h S c h o o l of Medicine, PHS
Grant
Scientist D e v e l o p m e n t NIMH
NS-28512
Award,
and a Research
KO2-MH-00880
from
to S G B .
1 Andrade, R. and Nicoll, R.A., Pharmacologically distinct actions of serotonin on single pyramidal neurones of the rat hippocampus recorded in vitro, J. Physiol., 394 (1987) 99-124. 2 Azmitia, E.C. and Segal, M., An autoradiographic analysis of the differential ascending projections of the dorsal and median raphe nuclei in the rat, J. Comp. Neurol., 179 (1978) 641-668. 3 Beck, S.G., 5-Carboxyamidotryptamine mimics only the 5-HT elicited hyperpolarization of hippocampal pyramidal cells via 5-HTtA receptor, Neurosci. Lett., 99 (1989) 101-106. 4 Beck, S.G. and Halloran, P.H., lmipramine alters fl-adrenergic, but not serotonergic, mediated responses in rat hippocampal pyramidal cells, Brain Res., 504 (1989) 72-81. 5 Chaput, Y., Blier, P. and deMontigny, C., In vivo electrophysiological evidence for the regulatory role ofautoreceptors on serotonergic terminals, J. Neurosci., 6 (1986) 2796- 2801. 6 Colino, A. and Halliwell, J.V., Differential modulation of three separate K-conductances in hippocampal CAI neurons by serotonin, Nature, 328 (1987) 73 77. 7 deMontigny, C. and Aghajanian, G.K., Tricyclic antidepressants: long-term treatment increases responsivity of rat forebrain neurons to serotonin, Science, 202 (1978) 1303 1306.
8 Molliver, M.E., Serotonergic neuronal systems: what their anatomic organization tells us about function, J. Clin. Psychopharmacol., 7 (1987) 3S-23S. 90leskevich, S. and Descarries, L., Quantified distribution of the serotonin innervation in adult rat hippocampus, Neuroscience, 34 (1990) 19--33. 10 Otmakhov, N.A. and Bragin, A.G., Effects of norepinephrine and serotonin upon spontaneous activity and responses to mossy fiber stimulation of CA3 neurons in hippocampal slices, Brain Res., 253 (1982) 173-183. 11 Pazos, A. and Palacios, J.M., Quantitative autoradiographic mapping of serotonin receptors in the rat brain. I. Serotonin-1 receptors, Brain Res., 346 (1985) 205 230. 12 Penington, N.J. and Reiffenstein, R.J., Lack of effect of antagonists on serotonin-induced inhibition in rat hippocampus, Can. J. Physiol. Pharmacol., 64 (1986) 1413-1418. 13 Segal, M., Physiological and pharmacological evidence for a serotonergic projection to the hippocampus, Brain Res., 94 (1975) 115131. 14 Segal, M., 5-HT antagonists in rat hippocampus, Brain Res., 103 (1976) 161--166. 15 Segal, M., Regional differences in neuronal responses to 5-HT: intracellular studies in hippocampal slices, J. Physiol., 77 (1981) 373 -375. 16 Wilner, S.A., deMontigny, C., Desroches, J., Desjardins, P. and Suranyi-Cadotte, B.E., Autoradiographic quantification of serotonin~A receptors in rat brain following antidepressant drug treatment, Synapse, 4 (1989) 347 352.
