Keywords: Lumbricidae, Lumbricus; method; blood pulsation rate.

Earthworms provide a convenient means of studying circulation of the blood in invertebrates. Earthworms (Genus Lumbricus) are easily obtained most of the year, they can be examined without surgical intervention, and only the simplest equipment is required. The pulsatile movements of the dorsal longitudinal blood vessel in a posterior to anterior direction can be observed with little difficulty. The thermal sensitivity of the pulsation rate can be measured by immersing the earthworms in water, and altering the water temperature gradually; earthworms can live for fairly long periods in well aerated water. The usual method is to immerse the earthworms in water in a petri dish at room temperature; allow the worms to equilibrate for 20 minutes and then determine the pulsation rate by looking through a zoom stereo microscope at one particular section of the dorsal blood vessel. The water temperature is then lowered gradually to a lower temperature, usually 12 deg. C., by adding colder water; another equilibration period follows, and the pulsation rate is again determined, the procedure repeated at 4 deg. C intervals. Good lighting helps. A pair of students have been known to monitor up to four or five worms at one time. The dorsal blood vessel reaction to various drugs at different concentrations can also be studied by the water immersion method.

Research workers in other countries have found that over half the worms require little or no restraint, and most of the others require only a light touch of one or two fingers. It was also suggested that overly active worms can be excluded from the exercise (Hill, 1976). They have not met the calibre of New Zealand worms. My students and I have found that most of the worms are active, and do not remain quiescent under the light microscope, except for temperatures close to freezing point. Restraining the worms with two fingers of each hand was accepted by the worms as a challenge to wriggle out from between the fingers as soon as possible. Worms will also make a vertical roll, and turn the dorsal blood vessel out of sight.

To remedy this situation with minimal disturbance to the worms, Mr K. Bailey suggested the use of glass tubing to hold the worms stationary; I added the refinements of perforations for access of drugs and dissolved oxygen, and possible skirting of the glass at each end to keep the tubes off the bottom of the petri dish. Immature worms were used mostly, as the smaller 10cm length tubes fitted inside large petri dishes. The tubes were designed to be a little larger than the worms in diameter, to allow easy access of the worms into the tubes, hold them without discomfort and for water circulation around the worms. An internal diameter of 2.0mm was mostly found to be too small for the worms to enter and they refused to enter the tubes. Worms could be persuaded to enter 3.0 mm tubes and the worms would crawl down the full length of the tubes. The distant opening was plugged with a twirl of cotton inserted for approximately 10mm page 9 length, with a cotton extrusion for withdrawal. After entry of the worm the near end was similarly plugged. Sideholes in the glass tubing had been made at regular intervals of 10mm. The holes were of 1mm diameter and the worms appeared to find it difficult to leave through these holes. I consider there should be a profusion of these small holes to allow easy entry of water and dissolved chemicals. 20 or 25mm of tubing at the head end of the worm can be without holes to discourage the worm from attempting to leave the tube. The worms sometimes double over in these 3.0mm internal diameter tubes.

Examination of the dorsal blood vessel can now be made at different angles to a lamp directed horizontally at the worm, with the tube immersed below water in a petri dish. The tube can be held lightly by the fingers, or better by forceps to prevent warmth from the fingers being transmitted to the worm. Rolling of the worm could be countered by rolling the tube in the opposite direction. In addition to the peristaltic movements of the dorsal blood vessel, white blood corpuscles can be seen moving through the general body cavity fluid.