Exp. Brain Res. 33,523-534 (1978)

Experimental Brain Research 9

Springer-Verlag 1978

Polysynaptic Activation of the Dentate Gyrus of the Hippocampal Formation: An Olfactory Input Via the Lateral Entorhinal Cortex* R.C. Wilson and O. Steward Departments of Neurosurgery and Physiology, University of Virginia School of Medicine, Charlottesville, VA 22901, U. S. A. Summary. The possibility that olfactory input is transmitted to specific subregions of the hippocampal formation via the entorhinal cortex was investigated electrophysiologically by analyzing the laminar profiles of potentials evoked in the hippocampal formation by stimulation of the lateral olfactory tract (LOT). L O T stimulation resulted in long latency (14-20 ms) evoked responses in the dentate gyrus of the hippocampal formation ipsilateral to the stimulation. The variable long latency of these responses and their inability to follow stimulus rates of 40/s suggested that these potentials reflected polysynaptic activation. Analysis of the laminar profiles of the evoked potentials indicated that the responses originated from a synaptic field localized in the outer portion of the stratum moleculare of the dentate gyrus, a terminal distribution which overlaps that of the lateral entorhinal cortical (LEC) projection to the dentate gyrus. Lesions of the L E C eliminated the long latency responses in the dentate gyrus evoked by L O T stimulation. In addition, a conditioning pulse delivered either to the L O T or to the L E C produced paired pulse potentiation of the response elicited by subsequent stimulation of the other structure. No evidence was found to indicate that responses were generated in regio superior of the hippocampus proper following L O T stimulation. Taken together, these results suggest that stimulation of the L O T activates the dentate gyrus of the hippocampal formation by multisynaptic pathways which relay through the lateral portion of the entorhinal area. This finding is discussed with regard to entorhinal cortical organization and the known olfactory projections to the LEC.

Key words: Lateral olfactory tract - Evoked potentials - Dentate gyrus Lateral entorhinal cortex. * Some of this material was presented in abstract form at the 7th Annual Meeting of the Societyfor Neuroscienee, 1977 Offprint requests to: O. Steward (address see above) 0 0 1 4 - 4 8 1 9 / 7 8 / 0 0 3 3 / 0 5 2 3 / $ 2.40

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The hippocampal formation was designated as a component of the olfactory brain by most early investigators (Broca, 1878; Elliot Smith, 1903). Cajal (1901), however, questioned whether olfactory-related fibers reached either the hippocampus or the entorhinal cortical area (which provides the main extrinsic innervation of the hippocampus). When experimental anatomical investigations also failed to demonstrate such connections (Fox and Schmidt, 1943; Le Gros Clark and Meyer, 1947), the olfactory hypothesis of hippocampal function fell into disfavor (Brodal, 1947). The possibility of a relationship between the hippocampus and the sense of smell was revived, however, when anatomical studies employing autoradiographic and improved degeneration methods revealed olfactory related projections to the entorhinal area. Fibers originating in the olfactory bulb were shown to terminate in the ventrolateral entorhinal cortex (LEC) (White, 1965; Heimer, 1968; Price, 1973). In addition, primary olfactory receiving areas such as the anterior prepyriform cortex (Cragg, 1961; Price, 1973) and periamygdaloid cortex (Krettek and Price, 1977) were observed to relay secondary projections to the same entorhinal region. That a direct projection from the olfactory bulb to the L E C exists was also suggested by electrophysiological studies (Kerr and Dennis, 1972). In view of these findings, a multisynaptic olfactory input to the hippocampal formation via the L E C seems probable and hippocampal evoked potentials following olfactory bulb stimulation have, indeed, been reported (Berry et al., 1952; Cragg, 1960). These previous studies did not, however, directly address possible entorhinal involvement in the olfactory responses or identify the activated subregions and synaptic fields within the hippocampal formation. These omissions are significant since a recent description of separate entorhinal projection systems to the dentate gyrus and to the regio superior division of the hippocampus proper (Steward, 1976; Steward and Scoville, 1976) raises the possibility that information may be selectively transmitted to these structures via different cell populations in the entorhinal cortex. The present study was therefore designed first, to determine whether stimulation of the lateral olfactory tract could activate the hippocampal formation via relays through the entorhinal cortex, and second, whether such activity involved entorhinal projections to the dentate gyrus, to regio superior, or to both.

Methods

Thirteen male Sprague-Dawley rats each weighing 250-400 g served as subjects in these experiments. Prior to surgery, each animal was anesthetized with chloralose-urethane (55 mg/kg and 0.2-0.4 g/kg, respectively) and anesthesia was maintained throughout the experiment by supplementary urethane injections as needed. The cortex overlyingthe hippocampaland entorhinal areas was exposed and covered with a pool of mineral oil and a small portion of the skull overlying the ipsilateral frontal eortex was removed to permit access to the lateral olfactory tract (LOT). A glass micropipette recording electrode (2-15 Mff~impedance) was stereotaxicallypositioned above the dentate gyrus of the hippocampal formation using bregma and midline references (4.0 P, 2.0 L) and lowered until granule cell injury discharges were detected. Bipolar stimulating electrodes (twisted strands of Belden No. 40 Teflon-coated stainless steel wire) were positioned stereotaxically above the LOT at the level of the anterior olfactory nucleus (5.0 A, 1.5 L) and above the medial and

Olfactory Input to the Hippocampal Formation

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lateral entorhinal cortices (1.5 anterior to the transverse sinus, 3.0 L; and 2.0 anterior to the sinus, 6.0 L, respectively, both at a 10~ angle away from the midline). The electrodes were then lowered to a position where stimulation evoked maximal responses in the dentate gyrus. Monophasic constant voltage squarewave pulses of 0.3 ms duration and 2-100 V amplitude were delivered to the entorhinal stimulating electrodes through an Ortec 4710 Dual Channel Stimulator and WPI stimulus isolation units. The LOT was stimulated by constant current biphasic squarewave pulses (0.3-0.5 ms, 1.0-5.0 mA) from a WPI Bipolar Stimulator. Evoked potentials were recorded with the aid of a WPI Model 701 Microprobe system, displayed on a Tektronix R5310 Storage Oscilloscope after filtering (0.1 Hz and 10 KHz) and stored on FM tape. The signals were also fed to a Nicolet Model 1072 Instrument Computer for signal averaging, and printed with a Moseley Model 2D-4, X-Y plotter. In one series of experiments involving three animals, after the stimulating electrode positions had been optimized as indicated by recordings in the dentate gyrus, the recording electrode was repositioned to detect regio superior responses to LOT and entorhinal cortical stimulation. In two other animals, lesions were placed sequentially in the medial and lateral entorhinal cortices by passing 0.5 mA direct current through the stimulating electrodes for 20 s. Upon completion of each experiment, the animal was deeply anesthetized with an overdose of urethane and transcardially perfused with a 0.9% saline-10% formalin solution. The brain was removed and divided in the coronal plane between the entorhinal and hippocampal electrode positions. The anterior portion of the brain was sectioned coronally and the posterior portion horizontally to optimize visualization of all electrode sites. The sections were processed histologically for cresyl violet staining and the positions of the electrodes and entorhinal lesions were identified.

Results A v e r a g e d e v o k e d p o t e n t i a l s r e c o r d e d in t h e d o r s a l b l a d e o f t h e d e n t a t e gyrus following s t i m u l a t i o n o f t h e l a t e r a l o l f a c t o r y t r a c t ( L O T ) a r e i l l u s t r a t e d in Figs. l a n d 2. T h e L O T e v o k e d r e s p o n s e c o n s i s t e d o f a n e g a t i v e l y d i r e c t e d w a v e in the d e n d r i t i c r e g i o n w h i c h r e v e r s e d to a p o s i t i v e p o t e n t i a l n e a r t h e g r a n u l e cell b o d y l a y e r (Fig. 2). L O T s t i m u l a t i o n e l i c i t e d such a r e s p o n s e in b o t h t h e d o r s a l a n d v e n t r a l b l a d e s o f t h e d e n t a t e gyrus. T h e r e s p o n s e o c c u r r e d with a l a t e n c y w h i c h v a r i e d f r o m 1 4 - 2 0 ms, failed to follow s t i m u l a t i o n r a t e s o f 4 0 / s (Fig. 1 A ) , a n d e x h i b i t e d an a p p a r e n t r e c r u i t m e n t effect ( G l o o r , 1955) in which r e p e t i t i v e s t i m u l a t i o n at r a t e s o f 0.5/s to 2/s p r o d u c e d g r a d u a l l y a u g m e n t i n g r e s p o n s e s (Fig. 1B). T h e v a r i a b i l i t y in o n s e t latency, t h e l o n g latency, a n d t h e i n a b i l i t y to follow r a p i d r a t e s o f s t i m u l a t i o n suggest t h a t t h e r e s p o n s e was e v o k e d polysynaptically. B e c a u s e o f t h e l a m i n a t e d o r g a n i z a t i o n o f t h e d e n t a t e gyrus, t h e l a m i n a r p r o f i l e o f e v o k e d p o t e n t i a l d i s t r i b u t i o n a l o n g t h e g r a n u l e cell d e n d r i t i c axis reflects t h e l o c a t i o n o f t h e a c t i v a t e d t e r m i n a l field ( G l o o r et al., 1963; L o m o , 1971a), a n d t h e site o f m a x i m a l e x t r a c e l l u l a r n e g a t i v i t y in t h e p r o f i l e c o r r e s p o n d s to t h e site o f t h e a c t i v a t e d s y n a p t i c p o p u l a t i o n . Thus, the n e g a t i v e r e s p o n s e in t h e d e n d r i t i c r e g i o n o f t h e d e n t a t e gyrus e v o k e d b y L O T s t i m u l a t i o n suggests t h e a c t i v a t i o n o f a s y n a p t i c field in t h e distal r e g i o n s o f the g r a n u l e cell d e n d r i t e s . T o c o m p a r e the l a m i n a r p r o f i l e o f e v o k e d p o t e n t i a l d i s t r i b u t i o n following L O T s t i m u l a t i o n with t h a t o b s e r v e d following s t i m u l a t i o n of t h e e n t o r h i n a l cortex, t h e L O T , l a t e r a l e n t o r h i n a l c o r t e x ( L E C ) , a n d m e d i a l e n t o r h i n a l c o r t e x ( M E C ) w e r e s t i m u l a t e d d u r i n g a single e l e c t r o d e pass t h r o u g h t h e d e n t a t e gyrus a n d h i p p o c a m p a l f o r m a t i o n .

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Polysynaptic activation of the dentate gyrus of the hippocampal formation: an olfactory input via the lateral entorhinal cortex.

Exp. Brain Res. 33,523-534 (1978) Experimental Brain Research 9 Springer-Verlag 1978 Polysynaptic Activation of the Dentate Gyrus of the Hippocampa...
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