NEUROTENSIN MODULATES SYNAPTIC TRANSMISSION IN THE NUCLEUS OF THE SOLITARY TRACT OF THE RAT W. N. OGAWA, b V. BAPTISTA, a J. F. AGUIAR a AND W. A. VARANDA a * a Department of Physiology, Faculty of Medicine of Ribeirão Preto/ USP, Av. Bandeirantes, 3900 14049-900 Ribeirão Preto, Brazil b University of Tocantins, Porto Nacional/TO, Brazil Abstract—Whole-cell patch clamp recordings were made from neurons of the rat subpostremal region of the nucleus tractus solitarius (NTS) in transverse brainstem slices. Neurotensin (NT) enhanced the firing rate of action poten- tials from 0.80.4 Hz in control to 1.91.3 Hz (n9) and increased their decay time. The peak amplitude of the after-hyperpolarization was decreased by 345% (n9). These effects were associated with a depolarization of 41 mV (n10) in the resting membrane potential and an in- crease in the input resistance (from 768220 M to 986220 M; n5) and were compensated by manually hyperpolarizing the cell to control values. In voltage clamp experiments NT decreased an outward current (from 488161 to 34096 pA at 40 mV; n5) which reversed near the potassium equilibrium potential. In addition, NT increased the frequency of both excitatory and inhibitory spontaneous synaptic currents, an effect blocked by tetro- dotoxin, and did not change the evoked excitatory or in- hibitory postsynaptic currents. The selective NTR1 recep- tor antagonist SR48692 reversibly blocked the effects of NT on both action potentials and spontaneous synaptic currents. Our results suggest that NTR1 receptors can modulate post-synaptic responses in neurons of the sub- postremal NTS by increasing cell excitability as a result of blockade of a potassium conductance. © 2004 IBRO. Pub- lished by Elsevier Ltd. All rights reserved. Key words: electrophysiology, slice, post-synaptic currents, excitability, depolarization. The nucleus tractus solitarius (NTS) has a strategic position in relation to the brainstem neuronal networks due to the multiple modalities of visceral sensory afferents that reaches it (Loewy, 1990). This variety of peripheral neural afferents and its extensive central connections, make the NTS a site containing heterogeneity and a vast assortment of neuro- transmitters/modulators and their receptors (Van Giersber- gen et al., 1992). Although a large number of neuroactive substances seem to be involved in the neurohumoral inte- gration within the NTS, in vitro electrophysiological record- ings have suggested that glutamate and GABA are the prin- cipal neurotransmitters released in response to electrical stimulation of afferent sensory inputs in several NTS subdi- visions (Mifflin and Felder, 1990; Brooks et al., 1992; Fortin and Champagnat, 1993; Andresen and Yang, 1995; Wang and Bradley, 1995; Smith et al., 1998). A considerable amount of experimental data suggests that the tridecapeptide neurotensin (NT) may play at least a modulator role in the processing of the viscerosensory information reaching the NTS (Kubo and Kihara, 1990; Ciriello and Zhang, 1997; Seagard et al., 2000). NT is a putative peptidergic neuro- transmitter (Stowe and Nemeroff, 1991; Hermans and Malo- teaux, 1998), first isolated and chemically characterized by Carraway and Leeman (1973) from hypothalamic extracts. NT receptors are present in neurons intrinsic to the NTS and in those sending projections to other CNS nuclei (Riche et al., 1990; Wang et al., 1992). Microinjections of NT intracere- broventricularly or directly into the NTS lead to a cardiovas- cular response involving components of the baroreflex (Rioux et al., 1981; Quirion et al., 1981; Sumners et al., 1981; Shido and Nagasaka, 1985; Kubo and Kihara, 1990; Ciriello and Zhang, 1997; Seagard et al., 2000). In this study we examine the electrophysiological con- sequences of activating NT receptors in neurons of the subpostremal NTS, in transverse brainstem slices using the whole cell patch clamp technique. We have found that NT leads to a depolarization of the neurons, by suppress- ing a potassium conductance, thereby increasing excitabil- ity of the system. EXPERIMENTAL PROCEDURES Brain slices Brainsteim slices were prepared from 20 to 25 days-old Wistar rats of either sex. Following decapitation and craniotomy, the brain and upper cervical spinal cord were removed and submerged in ice-cold (2–3 °C) artificial cerebrospinal fluid (ACSF) pH 7.35–7.4, equilibrated with 95% O 2 –5% CO 2 (carbogen). The ACSF con- tained (mM): 120 NaCl; 2.5 KCl; 1.0 MgCl 2 ; 2.0 CaCl 2 ; 25 NaHCO 3 ; 1.25 NaH 2 PO 4 ; 25 glucose and the osmolality was 305–310 mOsm/kg H 2 O. After the brain stem (approximately 8 mm) was dissected, it was glued with cyanoacrylate glue, through its ventral surface, to the vertical arm of an L-shaped agar block (4% agar in ACSF) and taken to the stage of a Vibratome (MA756; Campden Instruments, Leicester, UK). Routinely, two transverse slices (300 m thick) containing the subpostremal NTS were obtained from each animal. After cutting, the slices were incubated for 60 min at 32 °C with ACSF constantly gassed with carbogen. A single slice was transferred to the recording chamber (0.4 ml) on the stage of an upright microscope (E600; Nikon Inc., *Corresponding author. Tel: +55-16-602-3024; fax: +55-16-633- 0017. E-mail address: wvaranda@fmrp.usp.br (W. A. Varanda). Abbreviations: ACSF, artificial cerebrospinal fluid; AHP, after- hyperpolarization; AP5, D(-)-2-amino-5-phosphonopentanoic acid; DNQX, 6,7-dinitroquinolixaline-2,3 dione; NMDA, N-methyl-D- aspartate acid; NT, neurotensin; NTS, nucleus tractus solitarius; NTS1, high affinity neurotensin receptor; sEPSC, spontaneous ex- citatory postsynaptic currents; sIPSCs, spontaneous inhibitory postsynaptic currents; SR49682, non-peptide antagonist of the high affinity neurotensin receptor; ST, solitary tract; TTX, tetrodotoxin. Neuroscience 130 (2005) 309 –315 0306-4522/05$30.00+0.00 © 2004 IBRO. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.neuroscience.2004.09.019 309