Heightened nicotinic regulation of auditory cortex during adolescence

Abstract

Adolescent smoking is associated with auditory-cognitive deficits and structural alterations to auditory thalamocortical systems, suggesting that higher auditory function is vulnerable to nicotine exposure during adolescence. Although nicotinic acetylcholine receptors (nAChRs) regulate thalamocortical processing in adults, it is not known whether they regulate processing at earlier ages since their expression pattern changes throughout postnatal development. Here we investigate nicotinic regulation of tone-evoked current source density (CSD) profiles in mouse primary auditory cortex from just after hearing onset until adulthood. At the youngest ages, systemic nicotine did not affect CSD profiles. However, beginning in early adolescence nicotine enhanced characteristic frequency (CF)-evoked responses in layers 2-4 by enhancing thalamocortical, early intracortical, and late intracortical response components. Nicotinic responsiveness developed rapidly and peaked over the course of adolescence, then declined thereafter. Generally, responsiveness in females developed more quickly, peaked earlier, and declined more abruptly and fully than in males. In contrast to the enhancement of CF-evoked responses, nicotine suppressed shorter-latency intracortical responses to spectrally distant (non-CF) stimuli while enhancing longer-latency responses. Intracortical infusion of nAChR antagonists showed that enhancement of CF-evoked intracortical processing involves α4β2*, but not α7, nAChRs, whereas both receptor subtypes regulate non-CF-evoked late intracortical responses. Notably, antagonist effects in females implied regulation by endogenous acetylcholine. Thus, nicotinic regulation of cortical processing varies with age and sex, with peak effects during adolescence that may contribute to the vulnerability of adolescents to smoking.

Figure 1.

Tone-evoked CSD profiles in adolescent mouse A1. A, B, LFPs and derived CSD profiles evoked in response to CF (A; 22 kHz) and non-CF (B; 5.5 kHz; both 60 dB SPL) tones in a P32 female mouse. In CF-evoked CSD traces (top, middle column), the layer 3/4 sink (green trace) had the earliest onset in the upper layers, and the CF main sink (red trace) occurred 100 μm more superficially. Note that the onset of the layer 5/6 sink (blue trace) is earlier than that of the layer 3/4 sink. The non-CF main sink (bottom, middle column, orange trace) was 100 μm below the depth of the CF main sink and had a longer onset latency (vertical dotted lines are aligned to the onset of the CF-evoked layer 3/4 sink). Response traces are from recording sites separated by 100 μm, and trace duration also indicates tone duration (100 ms). Calibration: LFP, 0.2 mV; CSD, 10 mV/mm². CSD color contours (right) were constructed by normalizing CSD traces to maximum and minimum values; thick horizontal lines indicate five response phases: CF-evoked input (1), early intracortical (2), and late intracortical (3) phases, and non-CF-evoked early (4) and late (5) phases. Numbers on far left indicate cortical layer. Laminar proportions are based on adult values and are estimates only (see Materials and Methods for details). WM, White matter.

Figure 2.

A, B, Periadolescent development of tone-evoked current sink magnitudes (A) and onset latencies (B). A, Magnitudes of CF tone-evoked current sinks for input (black), early intracortical (white), and late intracortical (gray) phases for five age groups, normalized to magnitudes at P21–P25. Current sink areas for P21–P25 were 51.8, 1671, and 3865 mV · ms/mm² for input, early intracortical, and late intracortical phases, respectively. No significant differences were observed among age groups (p = 0.71, 0.97, and 0.93 for input, early intracortical, late intracortical phases, respectively; five-age, one-way ANOVA). B, Onset latencies of CF tone-evoked layer 3/4 sink (black), layer 5/6 sink (white), and the non-CF main sink (gray). Asterisks indicate differences between groups (p < 0.05, two-tailed t test).

Figure 3.

Effects of systemic nicotine on tone-evoked CSD profiles become apparent during adolescence. A, B, Nicotine had little effect on CSD profile in a preadolescent female (P22; CF stimulus, 21 kHz, 65 dB SPL) (A), but enhanced CSD profile in an early-adolescent female (P28; CF stimulus, 20 kHz, 65 dB SPL) (B). Each CSD profile is an average of three from within a 12 min time period: for example, before (Control), 0∼12 min after (compare with Fig. 4A, shaded area), 24∼36 min after nicotine (Nic), with profiles for each animal normalized to maximum source and sink values among all conditions.

Figure 4.

Nicotinic effects on CF-evoked CSD profiles peak during adolescence, earlier for females than males. A, Time course of nicotine's effect on the input phase of layer 3/4 sink. Current sink areas are normalized to prenicotine baseline in males (top left) and females (top right) for each age group: P21–P25 (•); P26–P30 (■); P31–P35 (□); P36–P40 (▴); >P70 (○). Vertical solid lines at 0 min indicate the time of nicotine injection. Error bars for some data omitted for clarity. Bottom, Time course of nicotine effect at each age, plotted for 12 min postnicotine intervals (vertical dotted lines at top): 0∼12 min (■); 12∼24 min (•); 24∼36 min (□); 36∼48 min (○). B, C, Time course of nicotine effect at each age for early intracortical (B) and late intracortical (C) phases of the CF main sinks. D, Time course at each age of nicotine effect on onset latencies of CF tone-evoked layer 3/4 sinks averaged; symbols are as given above, and latencies were normalized to prenicotine baseline. Error bars (±SEM) are indicated upward for black symbols and downward for white symbols for clarity. *Prenicotine versus postnicotine, p < 0.05, paired t test; ^§significant differences compared with P21–P25; ^¶significant differences compared with >P70 (p < 0.05, one-way ANOVA). The numbers by the symbols indicate the group time points with 1 for 0∼12 min, 2 for 12∼24 min, 3 for 24∼36 min, and 4 for 36∼48 min.

Figure 5.

Nicotine has a different effect on non-CF-evoked CSD profiles than on CF-evoked profiles. A, An example of nicotine's differential effects on CSD profiles evoked by CF vs non-CF stimuli (P30, female). Nicotine enhanced all three phases of the CF main sink, but for the non-CF main sink it reduced the early phase (labeled 4) while it enhanced the late phase (labeled 5). CSD profiles are averages of three time points within 12 min intervals before (Control), and 0∼12, 12∼24, 24∼36, and 36∼48 min after nicotine injection (Nic). Maximum and minimum values used for normalization were 50.7 and −51.9 mV/mm² for CF, and 11.2 and −17.5 mV/mm² for non-CF, respectively. B, Effect of nicotine on the early phase of non-CF main current sinks. Time course of effects for males (B, left) and females (B, right) for each age group: P21–P25 (•); P26–P30 (■); P31–P35 (□); P36–P40 (▴); >P70 (○). Error bars omitted for clarity. C, Average nicotine effect on early phase (first 12 min after nicotine, i.e., shaded area in B) for males (■) and females (□). D, Effect of nicotine on late phase; asterisk for data at P36–P40 applies to males only. Error bars: ±SEM. *Prenicotine vs postnicotine, p < 0.05, paired t test.

Figure 6.

Effects of intracortical DHβE injection on nicotinic regulation of CF tone-evoked current sinks. A–C, Time course of effects on Input (A), Early Intracortical (B), and Late Intracortical (C) current sinks are shown for males (P36–P40; left column) and females (P26–P30; middle column). ACSF (□) or DHβE (■; 1 μm at fluid port) was injected intracortically over 10 min (shaded area). Nicotine was injected systemically at the vertical lines (0 min). Right column, Mean current sink magnitude (average of 5 time points) before (□) and after (■) nicotine in the presence of ACSF (males, n = 6; females, n = 8) or DHβE (males, n = 5; females, n = 7) for males (left side) and females (right side). Error bars: ± SEM. *p < 0.05, before versus after nicotine, paired t test; ^§p < 0.05, baseline control versus DHβE, paired t test.

Figure 7.

Effects of intracortical MLA injection on nicotinic regulation of CF tone-evoked current sinks. A–C, Time course of effects on Input (A), Early Intracortical (B), and Late Intracortical (C) current sinks are shown for males (P36–P40; left column) and females (P26–P30; middle column). ACSF (□) or MLA (■, 10 nm at fluid port) was injected intracortically over 10 min (shaded area). Nicotine was injected systemically at the vertical lines (0 min). Right column, Mean current sink magnitude (average of 5 time points) before (□) and after (■) nicotine in the presence of ACSF (males, n = 6; females, n = 8) or MLA (males, n = 7; females, n = 6) for males (left side) and females (right side). Error bars: ±SEM. *p < 0.05, before versus after nicotine, paired t test.

Figure 8.

Model of information processing in A1 and its regulation by nicotine. A, CF stimuli activate auditory thalamic (MGv) neurons, which provide input to layers 3/4 in A1 (1). The thalamocortical input evokes intracolumn intracortical activity (2; column indicated by shaded area), followed by possibly intercolumn activity (4). Non-CF stimuli similarly activate thalamocortical input and subsequent intracolumn activity, which then rapidly propagates to the CF site via long-distance, “horizontal” projections (3). Horizontal projections also, though more slowly, recruit intercolumn activity (4), which may overlap partially, though not fully (see Discussion), with similar CF-elicited activity. B, In early-adolescent females, late-adolescent males, and adult males, nicotine enhances (1, 2, and 4), but suppresses or does not affect (3). Receptors containing α4β2, but not α7, nAChR subunits mediate enhancement of 1 (Kawai et al., 2007) and 2 (moreover, sensitivity to DHβE was qualitatively greater in females than in males) and CF-evoked activity (4). Both α4β2* and α7 nAChRs mediate enhancement of non-CF-evoked activity (4). The receptor mechanism underlying nicotinic suppression of 3 is not clear. C, In early-adolescent males and late-adolescent and adult females, nicotine enhances 1 and suppresses 3, but has little effect on 2 and 4.