Hypocretin and nicotine excite the same thalamocortical synapses in prefrontal cortex: correlation with improved attention in rat

Abstract

Thalamic projections to prefrontal cortex are important for executive aspects of attention. Using two-photon imaging in prefrontal brain slices, we show that nicotine and the wakefulness neuropeptide hypocretin (orexin) excite the same identified synapses of the thalamocortical arousal pathway within the prefrontal cortex. Although it is known that attention can be improved when nicotine is infused directly into the midlayer of the prefrontal cortex in the rat, the effects of hypocretin on attention are not known. The overlap in thalamocortical synapses excited by hypocretin and nicotine and the lack of direct postsynaptic effects prompted us to compare their effects on a sustained and divided attention task in the rat. Similar to nicotine, infusions of hypocretin-2 peptide into the prefrontal cortex significantly improved accuracy under high attentional demand without effects on other performance measures. We show for the first time that hypocretin can improve attentional processes relevant to executive functions of the prefrontal cortex.

Figures

Figure 1.

Electrophysiological effects of nicotine and hypocretin on sEPSCs in layer V pyramidal neurons of prefrontal cortex. A, Sweeps in voltage clamp showing sEPSCs under basal conditions (1), in the presence of two concentrations of nicotine [300 nm (2) and 10 μm (3)], after a 30 min washout (4), and in the presence of two concentrations of hypocretin-2 peptide [10 nm (5) and 100 nm (6)]. B, Bar graph showing sEPSC frequency for hypocretin-2 peptide and nicotine in varied order (mean ± SEM; n = 5 neurons), as well as SSR591813, a partial agonist of α4β2 nicotinic receptors (n = 5 neurons). C, The graph shows sEPSCs (in 10 s bins) from one pyramidal neuron in the presence of 0.5 μm atropine to block muscarinic receptors. Asterisks denote Y-tube application of acetylcholine (1 mm; 5 s). There was a 5 min washout period between the first and second Y-tube applications. Nicotine (50 nm) was then applied in the bath for 15 min before a final Y-tube application of acetylcholine. D, The bar graph shows sEPSC frequency (mean ± SEM; n = 5 neurons) elicited by Y-tube applications of acetylcholine under these conditions. Error bars represent SEM. Hcrt, Hypocretin; Nic, nicotine.

Figure 2.

Postsynaptic calcium transients elicited by nicotine and hypocretin at a single spine in apposition with a thalamocortical terminal. A, Images show spines on a tertiary dendritic branch of a layer V pyramidal neuron (1) under baseline conditions and (2) during a nicotine-elicited (10 μm) postsynaptic calcium transient at the spine in apparent apposition to a thalamocortical terminal (inset). Red and blue graph lines show the green/red (G/R) intensity ratio obtained from regions of interest on the spines, indicated by the red and blue arrows, respectively. B, Images show spines (3) 30 min after nicotine washout and (4) during a hypocretin-elicited (100 nm) postsynaptic calcium transient. The graph shows hypocretin-elicited changes in G/R intensity ratio at the same spines as above. Scale bar, 2 μm. C, Image showing (5) high calcium elicited by a burst of action potentials. Scale bar, 5 μm. The graph shows control series of images acquired without application of neurotransmitter or drug. The arrow indicates a brief depolarizing current injection used to elicit the burst of spikes. D, The bar graph at the bottom right shows peak Z-scores (mean ± SEM) for G/R ratio in spines with (n = 6 spines) and without (n = 31 spines) apparent thalamocortical (TC) appositions in response to nicotine, hypocretin, and the burst of spikes. Error bars represent SEM. Nic, Nicotine; Hcrt, hypocretin.

Figure 3.

Behavioral effects of bilateral infusions of nicotine or hypocretin into the medial prefrontal cortex on a task of sustained, divided attention. A, Schematic showing midlayer infusion sites in the medial prefrontal cortex for the nine rats included in the statistical analysis. B, Performance accuracy (mean ± SEM; n = 9 rats) at the 0.3 and 1.0 s stimulus durations after bilateral infusions of saline (S), two doses of hypocretin-2 peptide (H; 100 and 300 pmol), and nicotine (N; 17 nmol) into the medial prefrontal cortex. The effect of hypocretin was dose related. Nicotine and the higher dose of hypocretin (300 pmol) each significantly increased accuracy at the most challenging stimulus interval (*p < 0.01). Error bars represent SEM.