The effects of nicotinic and muscarinic receptor activation on patch-clamped cells in the optic tectum of Rana pipiens

Abstract

Both nicotinic and muscarinic cholinergic receptors are present in the optic tectum. To begin to understand how the activation of these receptors affects visual activity patterns, we have determined the types of physiological responses induced by their activation. Using tectal brain slices from the leopard frog, we found that application of nicotine (100 microM) evoked long-lasting responses in 60% of patch-clamped tectal cells. Thirty percent of these responses consisted of an increase in spontaneous postsynaptic currents (sPSCs) and had both a glutamatergic and GABAergic component as determined by the use of 6-cyano-7-nitroquinoxaline-2,3-dione (50 microM) and bicuculline (25 microM), respectively. Remaining response types consisted of an inward membrane current (16%) and an increase in sPSCs combined with an inward membrane current (14%). All responses could be elicited in the presence of tetrodotoxin (0.5 microM). Muscarinic receptor-mediated responses, induced by carbachol (100 microM) application after nicotinic receptor desensitization, produced responses in 70% of tectal cells. In contrast to responses elicited by nicotine, carbachol-induced responses could be evoked multiple times without significant decrement. Responses consisted of either an outward current (57%), a decrease in sPSCs (5%) or an increase in sPSCs, with (almost 6%) or without (almost 3%) an outward current. The response elicited by carbachol was not predicted by the response of the cell to nicotine. Our results suggest that nicotinic receptors are found predominantly at presynaptic locations in the optic tectum while muscarinic receptors are most often present at postsynaptic sites. We conclude that both of these receptor types could substantially modulate visual activity by changing either the input to tectal neurons or the level of their response to that input.

Fig. 1

Responses produced in tectal cells by nicotinic receptor activation. One of three responses was evoked in tectal cells by application of nicotine (nic; 100 μM): an increase in the frequency and amplitude of sPSCs (A1, 30% of total recordings), an increase in the frequency and amplitude of sPSCs and an inward membrane current (B1, 14% of total recordings) or just an inward membrane current (C1, 16% of total recordings). Nicotine did not evoke any response in 40% of the cells. Arrows in A1, B1 and C1 indicate areas of traces displayed at a faster time scale prior to drug application (A2, B2 and C2) and during the peak period of the nicotine response (A3, B3 and C3).

Fig. 2

The increased sPSC responses induced by nicotine consist of glutamatergic and GABAergic activity. Substantial portions of the nicotine-induced increases in sPSCs were blocked by the non-NMDA receptor antagonist CNQX (50 μM; middle trace) and the GABA_A receptor antagonist bicuculline (25 μM; bottom trace). The solid line indicates the time during which nicotine was applied; the dotted line the time during which either CNQX or bicuculline was present. CNQX or bicuculline were introduced 30 s before the agonist (not shown). All recordings are from different cells. Abbreviations: nic, nicotine; bicu, bicuculline.

Fig. 3

Responses in tectal cells evoked by muscarinic receptor activation. Application of carbachol (cch; 100 μM), after desensitization of nicotinic ACL receptors, produced four different responses: an outward membrane current (A1, 57% of the cells), an increase in the frequency and amplitude of sPSCs superimposed upon an outward current (B1, 6% of total recordings), a decrease in the frequency and amplitude of sPSCs (C1, 5% of total recordings) or an increase in the frequency and amplitude of sPSCs (D1, 3% of total recordings). Carbachol did not induce any response in 30% of the cells. Arrows in A1, B1 and C1 indicate areas of traces displayed at a faster time scale prior to drug application (A2, B2 and C2) and during the peak period of the carbachol response (A3, B3 and C3).

Fig. 4

Repeatability of carbachol-induced outward-current responses. (A) Sequential 30-s applications of carbachol (cch; 100 μM) produced repeated outward-current responses. Top trace is the response produced upon the initial application of carbachol; bottom trace, the response on the fourth application. The interval between carbachol applications is 5 min. (B) Amplitude and/or frequency, latency and duration of sequential responses to carbachol are plotted as a percent of the response to the initial application (trial 1). Carbachol-induced outward-current response features are relatively stable but have a non-significant tendency to decrease in amplitude and total current, and increase in latency with increasing trial number (n=3).

Fig. 5

Atropine blocks carbachol-induced responses. (A) Carbachol (100 μM) application induced outward current in a tectal cell (top trace). A second application of carbachol in the presence of atropine (2 μM) elicited a minimal response in this same cell. (B) A carbachol-induced increase in sPSCs (top trace) was prevented by the presence of atropine (bottom trace). Solid line indicates the time during which carbachol (cch) was applied; dotted line, the time during atropine was present. The interval between the beginning of the first and second applications of carbachol in both A and B was 5 min.