RESPONSE OF THE FROG OLFACTORY SYSTEM 185 needed for a steady state to be reached. The apparatus may therefore be used for experiments which have not, it seems, been possible before. One of its first uses has been in the investigation of the relationship between stimulus concentration and the amplitude of the resulting EOG. METHODS The animals and their preparation Common frogs (Rana temporaria) were used for the EOG recordings. A frog was first anaesthetized by placing it in 15 ml of 10• aqueous urethane in a beaker. As soon as the frog lost its reflexes, it was rinsed with water and placed in a holder. The frog remained anaesthetized by this treatment for the whole course of the experiment and it was not allowed to recover. The olfactory epithelium was exposed by dissecting away the dorsal wall of the nasal cavity opposite to the eminentia olfactoria. The opening thus made was 2-3 mm across and provided access for the recording electrode and the stream of air carrying the stimulus. The stimulation system The stimulation system may best be described by reference to Fig. 2. It consists of a constant stream of clean moist air which plays on the olfactory epithelium all the time. This prevents the mucus from drying up, prevents Flow Rotary regulator flowmeter Carrier (100mMmln) pressure Pressure Flow L ............ Water/ice bath Regulators gauges RestrictorS Odorant tubes Figure 2. Diagram of the arrangement of odorant streams.

186 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS extraneous odours and, when required, carries the odour stimulus to the sensory area. This stream flows at 50 ml min -• and emerges from a jet 1 mm inside diameter in laminar flow. (Mean linear velocity--100 cm s -• approx.) The six odorant streams are switched into the carrier stream as close to the point of discharge as possible so that downstream there is minimum dead space and wall surface to delay delivery. The volume of the dead space is in fact about 0.15 ml so that with a flow rate of 50 ml min -• there would be a delay of 0.18 s before a stimulus reached 63•o (-- 1 - l/e) of its full strength. This would represent the worst possible case, i.e. instantaneous forward mixing in the nozzle causing 'rounding' of the stimulus profile. (If there were no forward mixing in the nozzle, the stimulus would still take 0.18 s to reach the orifice but would arrive there at full strength.) The odorant streams (up to six in number) are generated by passing clean dry air over pools of liquid odorants held in U tubes. These U tubes have a straight central portion so that the air stream passes over about 8 cm a liquid surface without bubbling through it. This prevents formation of spray which might be carried forward and upset the concentration. The air flow through each tube can be regulated from about 1 ml min -• to 5 ml min -• by controlling the pressure to a sintered stainless steel flow restrictor before the U tube--or up to 10 ml rain -• by changing the flow restrictor. At these small flow rates the vapour leaving the U tube is practically in equilibrium with the liquid odorant. If necessary the U tubes can be im- mersed in a water bath to keep their temperature constant at or below room temperature. The odorants used are chemicals whose purity has been checked by glc analysis of head-space samples. They are used either neat or diluted with water or paraffin oil. (The paraffin oil used is first deodorized by treatment with activated silica.) This dilution is the means most used to provide widely different rates of delivery of odorant. The rates can also be regulated by adjusting the air flow rates or by cooling the U tubes in order to lower the vapour pressures. The odorants are conveyed to the applicator (where they are switched into the main carrier stream) by means of PTFE tubing of inside diameter 0.4 mm. This tubing is conveniently flexible and is easily and cheaply replaceable. It does absorb some of the odorant but, some minutes after starting the flow, it reaches a steady state which is not disturbed by the switching operation since the flow is not thereby interrupted. It is an important feature of the design of the system that this should be so. All