Calcium dependence of depolarization-induced suppression of inhibition in rat hippocampal CA1 pyramidal neurons

We made whole-cell recordings from CA1 pyramidal cells in the rat hippocampal slice preparation to study the calcium (Ca2+) dependence of depolarization-induced suppression of inhibition (DSI). DSI is a retrograde signalling process in which voltage-dependent Ca2+ influx into a pyramidal cell leads to a transient decrease in the release of GABA from interneurons. To investigate the Ca2+ dependence of DSI without altering extracellular divalent cations, we varied the type and amount of Ca2+ chelator (EGTA or BAPTA) in the recording pipette (keeping the chelator: Ca2+ ratio constant). Evoked inhibitory postsynaptic currents (IPSCs) were induced in the presence of antagonists of ionotropic glutamate receptors. DSI was induced by depolarizing voltage steps, lasting from 0·025 to 5 s, to 0 mV. DSI was directly dependent on the duration of the voltage step used to induce it, from threshold up to a maximal value of IPSC suppression, whether EGTA or BAPTA was used, and whether their concentrations were 0·1, 0·5 or 2 mM. For instance, a voltage step lasting 1·37 s produced half-maximal DSI with 2 mM BAPTA, but with 0·1 mM BAPTA, half-maximal DSI was achieved with a step lasting 0·186 s. Peak DSI was the same in all cases, and DSI was blocked with either 10 mM EGTA or BAPTA in the pipette. Bath application of carbachol could overcome the block of DSI by 10 mM EGTA but not by 10 mM BAPTA. We calculated that a voltage step lasting ≈100 ms would be necessary to activate half-maximal DSI in the absence of exogenous Ca2+ buffers. Log-log plots of calculated total Ca2+ influx, estimated from time integrals of Ca2+ currents, versus DSI yielded a straight line with a slope of ≈1, and increasing extracellular [Ca2+] from 2·5 to 5 mM did not change the slope. The time course of decay of DSI was well described by an exponential function with a time constant of ≈20 s and was not affected by changes in either concentration or type of Ca2+ buffer. The data suggest that, in its Ca2+ dependence, DSI more closely resembles the slow release of neuropeptides and hormones than it does the process of fast release of many neurotransmitters. At fast synapses, neurotransmission is tightly coupled to calcium influx and steeply dependent on the instantaneous internal [Ca2+] near the plasma membrane, which rises to high levels, and falls quickly with the opening and closing of voltage-dependent Ca2+ channels (VDCCs) (Dodge & Rahamimoff, 1967; Adler et al. 1991; Llinas et al. 1992; Borst et al. 1995; Borst & Sakmann, 1996; Wu & Saggau, 1997). However, chemical secretion is not always Ca2+ dependent in this way. Slow transmitter release from peptidergic nerve terminals (Peng & Zucker, 1993; Seward et al. 1995) or chromaffin cells (Neher & Zucker, 1993; Chow et al. 1994; Engisch & Nowycky, 1996; Seward & Nowycky, 1996) is typically initiated by prolonged Ca2+ influx and appears to be dependent on the [Ca2+] at a substantial distance from the VDCCs, rather than on the instantaneous submembrane [Ca2+]. Principal cells in the brain can release chemicals, often from dendrites, that travel in the retrograde direction and influence incoming signals. Several unconventional modulators, for example nitric oxide and carbon monoxide (Dawson & Snyder, 1994), may be retrograde messengers, but conventional neurotransmitters may also act in this way. Examples include dopamine (Cheramy et al. 1981; Jaffe et al. 1998) in the substantia nigra, dynorphin in the dentate gyrus (Drake et al. 1994), and oxytocin and vasopressin in the supraoptic nucleus (Kombian et al. 1997). By affecting cells synapsing on it, a cell releasing a retrograde messenger could affect its own state of excitability. Although retrograde signalling is triggered by Ca2+ influx, there is almost no information on its Ca2+ dependence. A well-established retrograde signal process exists in the hippocampus (Pitler & Alger, 1992, 1994; Alger & Pitler, 1995; Alger et al. 1996; Morishita & Alger, 1997; Ohno-Shosaku et al. 1998) and cerebellum (Llano et al. 1991; Vincent et al. 1992; Vincent & Marty, 1993) whereby voltage-dependent Ca2+ influx into a principal cell causes a transient reduction in the release of GABA from interneurons. This process, called depolarization-induced suppression of inhibition (DSI), is probably mediated by release of a messenger, perhaps glutamate or a glutamate analogue (Glitsch et al. 1996; Morishita et al. 1998), from somato-dendritic regions of the principal cells. DSI thus provides a robust model for investigation of retrograde neurotransmission. DSI could involve an unusual release mechanism. It is experimentally induced by long cellular depolarizations, usually > 1 s, and has a slow onset and long duration, typically not achieving peak levels for several seconds following postsynaptic depolarization and lasting for 20–60 s at 30°C. Hence, DSI could require large and prolonged increases in [Ca2+]i and have slow intrinsic kinetics. However, the dynamics of [Ca2+]i changes depend not only on the spatiotemporal pattern of Ca2+ influx, but also on endogenous and exogenous Ca2+ buffers (Zhou & Neher, 1993). The calcium buffers, BAPTA or EGTA, used at millimolar concentrations in experiments on DSI, could have affected its properties. By varying the intracellular Ca2+ buffering conditions, we have begun to explore the Ca2+ dependence of DSI in CA1 pyramidal cells. This approach, extensively used in investigations of neurotransmitter release (Adler et al. 1991; von Gersdorff & Matthews, 1994, 1997; Borst et al. 1995; Borst & Sakmann, 1996; Mennerick & Matthews, 1996), is particularly suitable for DSI because it directly affects [Ca2+]i without influencing other cells or the extracellular environment. The data suggest that DSI more closely resembles hormone or peptide secretion than it does fast synaptic neurotransmission.